Automated analysis of single cells using Laser Tweezers Raman Spectroscopy.

In recent years, significant progress has been made into the label-free detection and discrimination of individual cancer cells using Laser Tweezers Raman Spectroscopy (LTRS). However, the majority of examples reported have involved manual trapping of cells, which is time consuming and may lead to different cell lines being analysed in discrete batches. A simple, low-cost microfluidic flow chamber is introduced which allows single cells to be optically trapped and analysed in an automated fashion, greatly reducing the level of operator input required. Two implementations of the flow chamber are discussed here; a basic single-channel device in which the fluid velocity is controlled manually, and a dual-channel device which permits the automated capture and analysis of multiple cell lines with no operator input. Results are presented for the discrimination of live epithelial prostate cells and lymphocytes, together with a consideration of the consequences of traditional 'batch analysis' typically used for LTRS of live cells.

Car Safety Seat Usage and Selection Among Families Attending University Hospital Limerick.

The safest way for children to travel within a car is by provision of a weight-appropriate safety-seat. To investigate this, we conducted a cross-sectional study of adult parents who had children under 12 years, and collected information related to: car use, safety-seat legislation, and type of safety-seat employed. Data were reviewed on 120 children from 60 respondents. Ninety-eight (81.7%) children were transported daily by car. Forty-eight (81.4%) respondents were aware that current safety-seat legislation is based on the weight of the child. One hundred and seven (89.9%) children were restrained during travel using a car safety-seat. One hundred and two (96.2%) safety seats were newly purchased, installed in 82.3% (88) cases by family members with installation instructions fully read in 58 (55.2%) cases. Ninety-nine (83.2%) children were restrained using an appropriate safety-seat for their weight. The results show that four out of five families are employing the most appropriate safety-seat for their child, so providing an effective mechanism to reduce car-related injury. However, the majority of safety-seats are installed by family members, which may have child safety consequences.

The Establishment of a Pilot Paediatric Obesity Clinic at the University Hospital, Limerick.

This study describes the establishment of a pilot Paediatric Obesity Clinic (POC) in the University Hospital Limerick (UHL). Referrals were received from consultant paediatricians in the catchment areas of UHL for paediatric patients with high levels of excess adiposity. Fifteen patients and their families were invited to the POC in 2012. An initial medical assessment was conducted by 2 consultant paediatricians. Patients were also reviewed by a dietitian, a physiotherapist and physical activity experts from local Sports Partnerships. Twelve children and their families attended the POC (mean age = 8.08 years; Range = 3.6-13.6): 11/12 were overweight and 9/12 were obese. Abnormalities in blood work were detected as follows: 1/7 had elevated LDL-cholesterol; 2/8 had elevated triglyceride levels; 4/8 had elevated fasting insulin; 2/8 had elevated fasting glucose. With the current prevalence of obesity in paediatric populations, initiatives such as UHL's POC need to be established, funded and supported, to try to meet complex, multidisciplinary patient needs and to prevent future complex and expensive health complications.

Transition to adult care for adolescents with diabetes--a national survey.

Currently, there are no national guidelines on transition from paediatric to adult services for children with Type 1 Diabetes Mellitus (T1DM) in Republic of Ireland. There are 19 hospitals in Republic of Ireland looking after children with T1DM. Seventeen have a designated clinic for children with T1DM. Ten have a transition clinic for adolescents with T1DM. Most centres transition after patients finish secondary education. Six centres hold transition information sessions and 6 have access to a psychologist. Fifteen centres describe a gradual transition process. There is little national consistency in transition and there is a need for a collaborative national framework on T1DM transition.

The microbiome-gut-brain axis during early life regulates the hippocampal serotonergic system in a sex-dependent manner.

Bacterial colonisation of the intestine has a major role in the post-natal development and maturation of the immune and endocrine systems. These processes are key factors underpinning central nervous system (CNS) signalling. Regulation of the microbiome-gut-brain axis is essential for maintaining homeostasis, including that of the CNS. However, there is a paucity of data pertaining to the influence of microbiome on the serotonergic system. Germ-free (GF) animals represent an effective preclinical tool to investigate such phenomena. Here we show that male GF animals have a significant elevation in the hippocampal concentration of 5-hydroxytryptamine and 5-hydroxyindoleacetic acid, its main metabolite, compared with conventionally colonised control animals. Moreover, this alteration is sex specific in contrast with the immunological and neuroendocrine effects which are evident in both sexes. Concentrations of tryptophan, the precursor of serotonin, are increased in the plasma of male GF animals, suggesting a humoral route through which the microbiota can influence CNS serotonergic neurotransmission. Interestingly, colonisation of the GF animals post weaning is insufficient to reverse the CNS neurochemical consequences in adulthood of an absent microbiota in early life despite the peripheral availability of tryptophan being restored to baseline values. In addition, reduced anxiety in GF animals is also normalised following restoration of the intestinal microbiota. These results demonstrate that CNS neurotransmission can be profoundly disturbed by the absence of a normal gut microbiota and that this aberrant neurochemical, but not behavioural, profile is resistant to restoration of a normal gut flora in later life.

Bifidobacterium infantis suppression of Peyer's patch MIP-1α and MIP-1ß secretion during Salmonella infection correlates with increased local CD4+CD25+ T cell numbers.

The outcome following infection depends on the generation of an immune response that results in control of the pathogenic microorganism, while limiting inflammatory collateral damage to the host. Bifidobacterium infantis 35624 was shown to be protective against Salmonella associated host injury via a Treg-dependent mechanism. In this study, we further examined the mechanisms by which B. infantis-induced Tregs protect against Salmonella-associated inflammation. B. infantis 35624 feeding to Salmonella-infected mice significantly reduced Peyer's patch MIP-1α and MIP-1ß secretion. Chemokine secretion was significantly inversely correlated with Peyer's patch CD4+CD25+ cell numbers. In vitro, CD25+ T cells, but not CD25- T cells, specifically inhibited TNF-α and IFN-γ secretion. However, both CD25+ and CD25- T cells suppressed MIP-1α and MIP-1ß secretion to the same extent. This study suggests that although B. infantis 35624-induced Tregs correlate with inhibition of chemokine secretion within the mucosa of pathogen infected animals, indirect cellular mechanisms may play a role.

Diagnostic trajectory, interplay and convergence/divergence across all 12 DSM-IV psychotic diagnoses: 6-year follow-up of the Cavan-Monaghan First Episode Psychosis Study (CAMFEPS).

BACKGROUND: The boundaries of psychotic illness and the extent to which operational diagnostic categories are distinct in the long term remain poorly understood. Clarification of these issues requires prospective evaluation of diagnostic trajectory, interplay and convergence/divergence across psychotic illness, without a priori diagnostic or other restrictions. METHOD: The Cavan-Monaghan First Episode Psychosis Study (CAMFEPS), conducted using methods to attain the closest approximation to epidemiological completeness, incepts all 12 DSM-IV psychotic diagnoses. In this study we applied methodologies to achieve diagnostic reassessments on follow-up, at a mean of 6.4 years after first presentation, for 196 (97%) of the first 202 cases, with quantification of prospective and retrospective consistency. RESULTS: Over 6 years, the 12 initial psychotic diagnoses were characterized by numerous transitions but only limited convergence towards a smaller number of more stable diagnostic nodes. In particular, for initial brief psychotic disorder (BrP), in 85% of cases this was the harbinger of long-term evolution to serious psychotic illness of diagnostic diversity; for initial major depressive disorder with psychotic features (MDDP), in 18% of cases this was associated with mortality of diverse causality; and for initial psychotic disorder not otherwise specified (PNOS), 31% of cases continued to defy DSM-IV criteria. CONCLUSIONS: CAMFEPS methodology revealed, on an individual case basis, a diversity of stabilities in, and transitions between, all 12 DSM-IV psychotic diagnoses over 6 years; thus, psychotic illness showed longitudinal disrespect to current nosology and may be better accommodated by a dimensional model. In particular, a first episode of BrP or MDDP may benefit from more vigorous, sustained interventions.

A study to assess smoking habits and smoking exposure in sportspeople.

BACKGROUND: Tobacco smoking is a leading public health concern and is the most preventable cause of morbidity and mortality worldwide. Sportspeople are no exception and those who smoke are predisposed to the same hazardous health effects as the general public, in addition to the potential effects it may have on their sporting performance. AIM: We aimed to ascertain the prevalence of tobacco consumption in a sporting population. We also endeavoured to quantify the use of electronic cigarettes (e-cigarettes) and assess exposure to passive smoking. DESIGN: Observational study. METHODS: A web-based e-questionnaire was distributed to participants from various sports across Ireland between November 2017 and January 2018, and data were analysed using SPSS. RESULTS: A total of 546 sportspeople completed the survey with more than twice as many male respondents. Of whom, 16% of participants were current smokers, with males significantly more likely to smoke (P < 0.001), 26% of rugby players were current smokers which was significantly higher when compared with other sports (P < 0.01), 10% of all participants were exposed to second-hand smoke for more than 1 h per day and 2% of all participants were current users of e-cigarettes. CONCLUSION: The prevalence of smoking in our study population was higher than other literature reports. Further studies are essential to evaluate the potential negative effects this may be having on sporting performance, career progression and indeed injury occurrence/rehabilitation. It is imperative to address the matter of smoking in athletes, not only for public health concerns but also considering they are important role models in our society.

Non-invasive assessment of ventriculo-arterial coupling using aortic wave intensity analysis combining central blood pressure and phase-contrast cardiovascular magnetic resonance.

BACKGROUND: Wave intensity analysis (WIA) in the aorta offers important clinical and mechanistic insight into ventriculo-arterial coupling, but is difficult to measure non-invasively. We performed WIA by combining standard cardiovascular magnetic resonance (CMR) flow-velocity and non-invasive central blood pressure (cBP) waveforms. METHODS AND RESULTS: Two hundred and six healthy volunteers (age range 21-73 years, 47% male) underwent sequential phase contrast CMR (Siemens Aera 1.5 T, 1.97 × 1.77 mm2, 9.2 ms temporal resolution) and supra-systolic oscillometric cBP measurement (200 Hz). Velocity (U) and central pressure (P) waveforms were aligned using the waveform foot, and local wave speed was calculated both from the PU-loop (c) and the sum of squares method (cSS). These were compared with CMR transit time derived aortic arch pulse wave velocity (PWVtt). Associations were examined using multivariable regression. The peak intensity of the initial compression wave, backward compression wave, and forward decompression wave were 69.5 ± 28, -6.6 ± 4.2, and 6.2 ± 2.5 × 104 W/m2/cycle2, respectively; reflection index was 0.10 ± 0.06. PWVtt correlated with c or cSS (r = 0.60 and 0.68, respectively, P < 0.01 for both). Increasing age decade and female sex were independently associated with decreased forward compression wave (-8.6 and -20.7 W/m2/cycle2, respectively, P < 0.01) and greater wave reflection index (0.02 and 0.03, respectively, P < 0.001). CONCLUSION: This novel non-invasive technique permits straightforward measurement of wave intensity at scale. Local wave speed showed good agreement with PWVtt, and correlation was stronger using the cSS than the PU-loop. Ageing and female sex were associated with poorer ventriculo-arterial coupling in healthy individuals.

Suppression of tumorigenicity of human prostate carcinoma cells by replacing a mutated RB gene.

Introduction of a normal retinoblastoma gene (RB) into retinoblastoma cells was previously shown to suppress several aspects of their neoplastic phenotype, including tumorigenicity in nude mice, thereby directly demonstrating a cancer suppression function of RB. To explore the possibility of a similar activity in a common adult tumor, RB expression was examined in three human prostate carcinoma cell lines. One of these, DU145, contained an abnormally small protein translated from an RB messenger RNA transcript that lacked 105 nucleotides encoded by exon 21. To assess the functional consequences of this mutation, normal RB expression was restored in DU145 cells by retrovirus-mediated gene transfer. Cells that maintained stable exogenous RB expression lost their ability to form tumors in nude mice, although their growth rate in culture was apparently unaltered. These results suggest that RB inactivation can play a significant role in the genesis of a common adult neoplasm and that restoration of normal RB-encoded protein in tumors could have clinical utility.

Inactivation of the retinoblastoma susceptibility gene in human breast cancers.

Mutational inactivation of the retinoblastoma susceptibility (RB) gene, a recessive cancer gene, has been implicated in the genesis of retinoblastoma and certain other human neoplasms. This gene is now shown to be inactivated in two of nine human breast cancer cell lines examined. The RB gene of one cell line had a homozygous internal duplication of a 5-kilobase region containing exons 5 and 6. The RB messenger RNA transcript was correspondingly lengthened, and its translation was probably terminated prematurely due to a shifted reading frame. The other cell line had a homozygous deletion of the RB gene that removed the entire gene beyond exon 2. The RB gene product, pp110RB, was not detectable in either cell line by immuno-precipitation with specific antibodies. These findings are significant in relation to proposed genetic mechanisms of breast cancer formation.

Kynurenine pathway in psychosis: evidence of increased tryptophan degradation.

The kynurenine pathway of tryptophan degradation may serve to integrate disparate abnormalities heretofore identified in research aiming to elucidate the complex aetiopathogenesis of psychotic disorders. Post-mortem brain tissue studies have reported elevated kynurenine and kynurenic acid in the frontal cortex and upregulation of the first step of the pathway in the anterior cingulate cortex of individuals with schizophrenia. In this study, we examined kynurenine pathway activity by measuring tryptophan breakdown, a number of pathway metabolites and interferon gamma (IFN-gamma), which is the preferential activator of the first-step enzyme, indoleamine dioxygenase (IDO), in the plasma of patients with major psychotic disorder. Plasma tryptophan, kynurenine pathway metabolites were measured using high-performance liquid chromatography (HPLC) in 34 patients with a diagnosis on the psychotic spectrum (schizophrenia or schizoaffective disorder) and in 36 healthy control subjects. IFN-gamma was measured using enzyme-linked immunosorbent assay (ELISA). The mean tryptophan breakdown index (kynurenine/tryptophan) was significantly higher in the patient group compared with controls (P < 0.05). IFN-gamma measures did not differ between groups (P = 0.23). No relationship was found between measures of psychopathology, symptom severity and activity in the first step in the pathway. A modest correlation was established between the tryptophan breakdown index and illness duration. These results provide evidence for kynurenine pathway upregulation, specifically involving the first enzymatic step, in patients with major psychotic disorder. Increased tryptophan degradation in psychoses may have potential consequences for the treatment of these disorders by informing the development of novel therapeutic compounds.

The relationship between gender and age with monocyte tissue factor expression.

BACKGROUND: In normal healthy individuals, the level of tissue factor (TF) expression on monocytes is low. However, studies have shown that patients with cardiovascular disease (CVD) have elevated levels of TF. As the risk of CVD increases with age and is more prominent in the male population, it is postulated that TF expression may be positively correlated with these factors. However, very few studies have examined the relationship between age and gender on TF expression. METHODS: This study evaluated the influence of age and gender on TF expression using data obtained from female (n = 44) and male (n = 27) subjects. We also examined the influence of BMI and total fat intake on TF expression in the same subjects. RESULTS: The results of our study found no significant difference in TF expression between the male and female subgroups. No correlation was found between TF and age, BMI or total fat intake in the male or female groupings. CONCLUSION: It may be postulated that the risk of CVD development in such populations may not be due to increases in TF expression with increasing age or gender differences.

Anion complexation via C-H...X interactions using a palladacyclic receptor.

A novel organometallic receptor binds anions in solution and in the solid state, with complexes stabilised through a series of C-HX interactions, as evidenced by 1H NMR spectroscopy, X-ray crystallography and computational models.

Migration of intraperitoneally injected thyroid cells in the Amazon molly, Poecilia formosa.

We have previously reported the development of an extensive invasive growth of the thyroid gland of the gynogenetic teleost, Poecilia formosa (the Amazon molly), following i.p. injection of UV- or gamma-irradiated thyroid cells. This result was surprising by comparison with mammalian work, in which the thyroid is rarely the site for tumor metastases, but the anatomy of the circulation of fish is different from mammals, and in fish the gills and thyroid gland would be among the first tissues in which injected cells might be arrested. Techniques using a fluorescent dye, 125I membrane label, or [3H]thymidine label were used to follow the distribution of i.p. injected cells in the Amazon molly. Fish sampled as soon as 30 min after injection had some labeled cells dispersed in the connective tissue around the ventral aorta and in the bases of the gills, and by 1 to 4 hr large numbers of cells had moved into the thyroid region. A few cells still persisted there 200 hr later. Experiments on the distribution of heat-killed cells indicated that the initial distribution of the cells was largely governed by mechanical factors. Injected cells would appear to be disseminated in fish by mechanisms similar to those in mammals.

Expression of the retinoblastoma gene product in bladder carcinoma cells associates with a low frequency of tumor formation.

Upon inactivation of both alleles of the retinoblastoma gene (RB), individuals develop the intraocular eye tumor, retinoblastoma. The gene encodes a Mr 110,000 phosphorylated nuclear protein that may be involved in regulation of the cell cycle. Besides retinoblastoma, mutations of the gene have been detected in several other types of tumors, including bladder carcinoma. Up to one-third of bladder carcinomas may contain mutations of the RB gene. Introducing the retinoblastoma gene into single retinoblastoma, osteosarcoma, or prostate carcinoma cell lines suppresses their tumorigenicity as assayed in nude mice. We have sought to extend these results by introducing the retinoblastoma gene into multiple bladder carcinoma lines, and analyzing several of the resulting, cloned lines. We have found that inhibition of tumorigenicity, as assayed by tumor growth in nude mice or growth of cells in soft agar, is the only consistent phenotype observed upon re-expression of RB in all bladder carcinoma cells examined. The effect of RB expression on growth and cellular morphology varied depending on the particular parental cell line. We conclude that RB expression generally correlates with reduced tumorigenicity, but not reduced growth rate, in bladder carcinoma cells.

Older patient hospital admissions following primary care referral: the truth is in the referring.

BACKGROUND: Health information has a major role in the planning of future healthcare provision. With current reconfiguration and cost saving measures, further demands are being placed on acute hospitals. AIM: To examine the elderly admissions and the referral documentation of older patients admitted to a tertiary level hospital. METHODS: A retrospective analysis of primary care referral documentation for all acute admissions of patients over 75 years to University Hospital Limerick (UHL) over a 2-month period. Documentation was analysed on the basis of patient demographics, presenting complaint and referral source. Primary care referral documentation was then analysed on the basis of presenting complaint, patient demographics, referrer details, and the clinical information provided. RESULTS: Over the 2-month period there were a total of 381 elderly admissions through the Emergency department. The most common presenting complaint was with shortness of breath (21.5 %). 42.5 % of admissions were from a primary care setting. 31.1 % of referrals were typed and 47.0 % handwritten. Over 90 % of referrals contained the patient's name, date of birth and address. 98.7 % of referrals included a presenting complaint and 54 % included a past medical history. 20 % of referrals listed known drug allergies, while 9.3 % documented social history or baseline functional status. Referral letters from general practice and after-hour services were largely similar. CONCLUSIONS: Almost all primary care referrals included the required details as per recent HIQA guidelines. The further inclusion of optional information relating to patient social or functional status, which are of particular relevance to the older population may help patient management.