Immortalization and characterization of human cell lines with mast cell and monocytic properties

Summary. We have previously derived a cell strain which had both mast cell and monocytic properties from the bone marrow of a child with diffuse cutaneous mastocytosis. This cell strain, termed HBM-M, consisted of two cell populations both of which possessed certain ultrastructural, cytochemi-cal and surface phenotypic features of degranulated mast cells. The cells also displayed cytochemical and surface phenotypic features of monocytes. These cells may represent a common bone marrow derived mast cell/monocyte precursor. Studies of human mast cells have been hindered by the fact that it is difficult to establish such cells in long-term culture. Thus, we sought to immortalize HBM-M cells by introducing Simian virus 40 large T-antigen. Following transfection by the strontium phosphate technique, transformed cells were selected, expanded and passaged until the cells entered a non-proliferative phase termed crisis. Certain clones passed through crisis 3 months later and by this means two immortal cell lines, HBM-MI-1 and HBM-MI-2, were obtained. The criterion for immortality was growth for greater than 100 population doublings. The immortal cell lines retained some, but not all. of the mast cell and monocytic properties of the original HBM-M cell strain. The immortalization of the cell strain HBM-M provides an opportunity to investigate the mast cell and monocytic properties of these cells, and the apparent relationship between mast cells and monocytes.     

Persistence of atovaquone in human sera following treatment: inhibition of Plasmodium falciparum development in vivo and in vitro

Published pharmacokinetic data indicate that after treatment of patients with therapeutic doses of atovaquone/proguanil hydrochloride (Malarone, GlaxoSmithKline Research Triangle Park, NC), the plasma half-lives of these drugs are 70h and 15h, respectively. However, using two biologic assays (mosquito transmission and in vitro asexual stage development), we demonstrate here that sera from volunteers treated with atovaquone/proguanil retained activity against Plasmodium falciparum up to 6 weeks after such treatment. This activity was due to atovaquone, as administration of this drug alone replicated the data obtained with the combination. Most notably, asexual stage development of an atovaquone-resistant strain (NGATV01) of P. falciparum was not inhibited by sera taken after atovaquone treatment. These data indicate that for atovaquone, biologic assays, though not quantitative, are more sensitive than the usual physicochemical assays. Also, persistence of atovaquone in plasma at low concentrations for long periods may increase the risk of resistant parasites arising.     

Radiographic hand joint space width assessed by computer is a sensitive measure of change in early rheumatoid arthritis

OBJECTIVE: To compare changes in the computerized measurement of radiographic hand joint space width (JSW) to changes in modified Sharp scores in a retrospective 2-year study of early rheumatoid arthritis (RA). METHODS: First and last standard clinical hand radiographs of 245 patients with RA were analyzed blind using purpose-written computer software to measure changes in JSW for proximal interphalangeal (PIP) and metacarpophalangeal (MCP) joints in the 3 middle fingers of each hand. Before measurement, the radiographs were scored independently by 2 radiologists using a modification of Sharp scoring. RESULTS: The paired changes in JSW (-0.051 +/- 0.005 mm) and Sharp score (+3.81 +/- 0.50) were both significant over the study duration. In measured joints showing an increase in joint space narrowing (JSN) score, 92% had a corresponding reduction in JSW. In patients with an increase in total score, including JSN and erosion scores in fingers and wrists, 84% had a corresponding reduction in mean (PIP + MCP) JSW. Patients with no change in Sharp score (47%) still experienced a significant reduction in measured JSW (-0.027 +/- 0.006 mm). HLA-DR genetic markers of severe disease progression were associated with significantly greater reductions in JSW but not increases in Sharp score. (Values: mean +/- standard error of mean). CONCLUSION: Measured JSW averaged over 6 PIP and 6 MCP joints was a valid and more sensitive measure of change than total Sharp score in this study of early RA.     

Respiratory virus infections in transplant recipients after reduced-intensity conditioning with Campath-1H: high incidence but low mortality

Respiratory virus infections can cause serious morbidity and mortality after conventional allogeneic stem cell transplantation. However, the incidence and outcome of these infections after reduced intensity conditioning has not been reported. Between 1997 and 2001, 35 episodes of respiratory virus infections were noted in 25 of 83 transplant recipients conditioned with fludarabine, melphalan and Campath-1H, and 80% of them received early antiviral therapy. Parainfluenza virus (PIV) 3 was the commonest isolate (45.7%) followed by respiratory syncytial virus (37%). Patients with myeloma were more susceptible to these infections [odds ratio (OR) 4.1, P = 0.01] which were often recurrent in patients with severe acute or chronic graft-versus-host disease (GVHD) (OR 10.6, P = 0.03). Infection within the first 100 d (OR 5.0, P = 0.05) and PIV 3 (OR 9.2, P = 0.01) isolation were risk factors for developing lower respiratory infection. Although more than half of the episodes progressed to lower respiratory infection, the mortality was only 8%. This could have been due to early initiation of antiviral therapy, but the attenuation of pulmonary damage due to the reduced-intensity conditioning, low incidence of GVHD and, paradoxically, the low CD4+ T-cell subset in this setting might also have been contributory factors.     

Low forced expiratory volume is associated with blunted cardiac reactions to acute psychological stress in a community sample of middle-aged men and women

It has been argued recently that blunted cardiovascular reactions to acute psychological stress have adverse behavioural and health corollaries that reflect dysregulation of the neural systems that support motivation. We examined the association between cardiovascular reactions to a standard stress task, the paced auditory serial arithmetic rest, and forced expiratory volume in one second, an effort, hence motivation, dependent assessment of lung function measured by spirometry. Low forced expiratory volume, expressed as a ratio to height squared was associated with blunted heart rate, but not blood pressure, stress reactivity, r = .17, p < .001. The association survived adjustment for smoking, a range of anthropometric and sociodemographic covariates, resting heart rate and stress task performance, β = .11, p = .005. As such, our results provide support for the hypothesis that blunted stress reactivity may be a peripheral marker of a dysfunction in the brain systems that support motivated behaviour.     

Evaluation of Irreversible Compression Ratios for Medical Images Thin Slice CT and Update of Canadian Association of Radiologists (CAR) Guidelines

In June 2008, the Canadian Association of Radiologists published its Standards for Irreversible Compression in Digital Diagnostic Imaging within Radiology (Canadian Association of Radiologists 2012). The study suggested that at low levels of compression there was no difference in diagnostic accuracy between uncompressed JPEG and JPEG 2000. There were two exceptions; CT neurological and CT body images resulted in lower rating of image quality (Koff et al., J Digit Imaging 22(6):569–78, 2009). The slice thicknesses used in the previous study were greater than 5 mm. However, other studies (Ringl et al., Radiology 240:869–87, 2006) suggest that thin CT slices might modify image tolerance to irreversible compression. Therefore, a new clinical evaluation using CT slices less than 3 mm was initiated. We examined CT images in four body regions (chest, body, musculoskeletal, and neurological). Twenty-five radiologists from across Canada participated. Each read a total of 70 CTs in his specialty; 10 at each of seven levels of compression (uncompressed, JPEG and JPEG 2000 at low, medium, and high compression (varying by region)). Each reader diagnosed the case, rated his confidence, and compared the compressed to the uncompressed image and rated the degree of degradation. Data were analyzed for sensitivity, specificity, accuracy, confidence, and degradation at three levels and two types of compression as well as the original image. There were no overall differences in sensitivity, specificity, accuracy, or confidence. JPEG images, at all levels of compression, were rated lower in terms of perceived difference (4.16/5 vs. 4.53/5 for JPEG 2000 and 4.68/5 for uncompressed). However, the rating of perceived difference was not significantly correlated with accuracy. Analysis of individual body regions did not reveal any systematic effects of compression in any region.     

Identifying Range-of-Motion Deficits and Talocrural Joint Laxity After an Acute Lateral Ankle Sprain

ContextApproximately 72% of patients with an ankle sprain report residual symptoms 6 to 18 months later. Although 44% of patients return to activity in less than 24 hours after experiencing a sprain, residual symptoms should be evaluated in the long term to determine if deficits exist. These residual symptoms may be due to the quality of ligament tissue and motion after injury.ObjectiveTo compare mechanical laxity of the talocrural joint and dorsiflexion range of motion (DFROM) over time (24 to 72 hours, 2 to 4 weeks, and 6 months) after an acute lateral ankle sprain (LAS).DesignCross-sectional study.SettingAthletic training research laboratory.Patients or Other ParticipantsA total of 108 volunteers were recruited. Fifty-five participants had an acute LAS and 53 participants were control individuals without a history of LAS.Main Outcome Measure(s)Mechanical laxity (talofibular interval and anterior talofibular ligament length) was measured in inversion (INV) and via the anterior drawer test. The weight-bearing lunge test was conducted and DFROM was measured. The data were analyzed using repeated-measures analysis of variance, independent-samples t tests, and 1-way analysis of variance.ResultsOf the 55 LASs, 21 (38%) were grade I, 27 (49%) were grade II, and 7 (13%) were grade III. Increases were noted in DFROM over time, between 24 and 72 hours, at 2 to 4 weeks, and at 6 months (P < .05). The DFROM was less in participants with grade III than grade I LASs (P = .004) at 24 to 72 hours; INV length was greater at 24 to 72 hours than at 2 to 4 weeks (P = .023) and at 6 months (P = .035) than at 24 to 72 hours. The anterior drawer length (P = .001) and INV talofibular interval (P = .004) were greater in the LAS group than in the control group at 6 months.ConclusionsDifferences in range of motion and laxity were evident among grades at various time points and may indicate different clinical responses after an LAS.     

Impaired glucose tolerance and reduced beta-cell function in overweight Latino children with a positive family history for type 2 diabetes

The objective of this study was to examine relationships between impaired glucose tolerance (IGT) and body composition and insulin-related phenotypes in 150 overweight Latino children with a family history of type 2 diabetes. Glucose tolerance was assessed by an oral glucose challenge. Body composition was assessed by dual energy x-ray absorptiometry and magnetic resonance imaging. Insulin sensitivity, the acute insulin response, and the disposition index (DI), as an index of beta-cell function, were determined by an iv glucose tolerance test and compared between normal glucose-tolerant and IGT children. IGT was present in 28% of children, and was similar across obesity groups, but higher in children exposed to gestational diabetes mellitus (41% IGT). There were no significant differences in body composition, fat distribution, insulin sensitivity, or acute insulin response, but DI was significantly lower in IGT children by 16% (P < 0.02), and DI was inversely related to age. In conclusion, IGT is present in 28% of overweight Latino children with a family history of type 2 diabetes, is not influenced by obesity, is more prevalent in children exposed to gestational diabetes mellitus, and is related to poor beta-cell function, which shows signs of deterioration with age in this population.     

Chromosome 15q25.1 genetic markers associated with level of response to alcohol in humans

As with other genetically complex common psychiatric and medical conditions, multiple genetic and environmental components contribute to alcohol use disorders (AUDs), which can confound attempts to identify genetic components. Intermediate phenotypes are often more closely correlated with underlying biology and have often proven invaluable in genetic studies. Level of response (LR) to alcohol is an intermediate phenotype for AUDs, and individuals with a low LR are at increased risk. A high rate of concurrent alcohol and nicotine use and dependence suggests that these conditions may share biochemical and genetic mechanisms. Genetic association studies indicate that a genetic locus, which includes the CHRNA5-CHRNA3-CHRNB4 gene cluster, plays a role in nicotine consumption and dependence. Genetic association with alcohol dependence was also recently shown. We show here that two of the markers from the nicotine studies also show an association (multiple testing corrected P < 0.025) with several LR phenotypes in a sample of 367 siblings. Additional markers in the region were analyzed and shown to be located in a 250-kb expanse of high linkage disequilibrium containing three additional genes. These findings indicate that LR intermediate phenotypes have utility in genetic approaches to AUDs and will prove valuable in the identification of other genetic loci conferring susceptibility to AUDs.