Force spectroscopy of hepatocytic extracellular matrix components.

We present atomic force microscopy and force spectroscopy data of live hepatocytes (HEPG2/C3A liver cell line) grown in Eagle's Minimum Essential Medium, a complex solution of salts and amino acids commonly used for cell culture. Contact-mode imaging and force spectroscopy of this system allowed correlation of cell morphology and extracellular matrix (ECM) properties with substrate properties. Force spectroscopy analysis of cellular "footprints" indicated that the cells secrete large polymers (e.g., 3.5mum contour length and estimated MW 1000kDa) onto their substrate surface. Although definitive identification of the polymers has not yet been achieved, fluorescent-labeled antibody staining has specified the presence of ECM proteins such as collagen and laminin in the cellular footprints. The stretched polymers appear to be much larger than single molecules of known ECM components, such as collagen and heparan sulfate proteoglycan, thus suggesting that the cells create larger entangled, macromolecular structures from smaller components. There is strong evidence which suggests that the composition of the ECM is greatly influenced by the hydrophobicity of the substrate surface, with preferential production and/or adsorption of larger macromolecules on hydrophobic surfaces.

Internal medicine residents reject "longer and gentler" training.

BACKGROUND: Increasing complexity of medical care, coupled with limits on resident work hours, has prompted consideration of extending Internal Medicine training. It is unclear whether further hour reductions and extension of training beyond the current duration of 3 years would be accepted by trainees. OBJECTIVE: We aimed to determine if further work-hour reductions and extension of training would be accepted by trainees and whether resident burnout affects their opinions. DESIGN: A postal survey was sent to all 143 Internal Medicine residents at the University of Colorado School of Medicine in May 2004. MEASUREMENTS: The survey contained questions related to opinions on work-hour limits using a 5-point Likert scale ranging from strongly agree to strongly disagree. Burnout was measured using the Maslach Burnout Inventory, organized into three subscales: emotional exhaustion (EE), depersonalization (DP), and personal accomplishment, with burnout defined as high EE or DP. RESULTS: Seventy-four percent (106/143) of residents returned the survey. The vast majority (84%) of residents disagreed or strongly disagreed with extending training to 4 or 5 years. Burnout residents were less averse to extending training (strongly agree or agree, 18.9% vs 4.3%, P = .04). The majority of residents (68.9%) disagreed or strongly disagreed with establishing a 60-hour/week limit. Residents who met the criteria for burnout were more likely to agree that a 60-hour limit would be better than an 80-hour limit (strongly agree or agree, 22% vs 8%, P = .02). CONCLUSIONS: In this program, most Internal Medicine residents are strongly opposed to extending their training to 4 or 5 years and would prefer the current 80 hours/week cap. A longer, less intense pace of Internal Medicine training seems to be less attractive in the eyes of current trainees.

Detection of Helicobacter pylori cagA gene in gastric biopsies, clinical isolates and faeces.

PURPOSE: Helicobacter pylori infection is common in the developing countries. The cagA gene is a marker of pathogenicity island (PAI) in H. pylori . The aim of this study was to determine the prevalence of cagA among dyspeptic patients in Bahrain directly from gastric biopsy and stool specimen. METHODS: A total of 100 gastric biopsy samples, 16 clinical isolates and 44 faecal specimens were collected from Bahraini adult dyspeptic patients. cagA gene of H. pylori was assessed using polymerase chain reaction (PCR). RESULTS: The cagA gene was detected in 59 (59%) from biopsy specimens, 10 (62%) clinical isolates and in 10 (22.7%) faecal specimens. The detection of cagA positive H. pylori was significantly higher in patients with duodenal ulcer (80%) compared to those with other endoscopic finding (42%) (P < 0.05). CONCLUSIONS: Using PCR to detect cagA gene directly from biopsy is a rapid and reliable technique. However, using stool specimen for genotyping in our patients showed reduced sensitivity.

Cocaine self-administration in rats differentially alters mRNA levels of the monoamine transporters and striatal neuropeptides.

The potential neuroadaptations to cocaine self-administration (SA) were evaluated using quantitative in situ hybridisation histochemistry. Levels of mRNAs of the monoamine transporters, i.e. the primary molecular targets of cocaine, and the striatal neuropeptides substance P and enkephalin, which predominantly exist in different populations of dopaminoceptive striatal neurons, were quantified in rats which had reached different stages of acquisition of cocaine SA. Thus, animals were killed 1 h after completing a self-administration session (i) early in or after acquisition of cocaine SA (ii) after various regimes of chronic cocaine SA, and (iii) a 10-day period of withdrawal from chronic cocaine intake. Control mRNA levels of all molecules under study were those quantified in animals receiving i.v. saline yoked to rats self-administering cocaine (1.5 or 0.75 mg/kg per infusion, depending on the experiment). Monoamine transporter expression was differentially altered by cocaine; dopamine transporter mRNA levels in the ventral tegmental area, but not in the substantia nigra, were increased following withdrawal from cocaine, suggesting a role for the upregulated mesolimbic dopamine transporter in the mechanisms underlying relapse to cocaine taking. By contrast, serotonin transporter mRNA in the dorsal raphé and noradrenaline transporter mRNA in the locus coeruleus remained unaltered under all experimental conditions. In addition, the expression of the striatal neuropeptides was also differentially altered; substance P mRNA levels were transiently increased in the shell of the nucleus accumbens by prolonged cocaine self-administration, but enkephalin mRNA levels in the dorsal and ventral striatum remained unaltered under all conditions.

Diagnosing coronary arterial stent thrombosis and arterial closure.

This single-center review of a consecutive series of patients requiring reexamination by angiography within 1 week of a coronary stent placement due to chest pain reveals that patients treated with a poststent anticoagulation regimen of warfarin and aspirin, and those with lower poststent deployment dilation pressures, have an increased risk of subacute stent thrombosis. Repeat cardiac catheterization within the first week after coronary artery stent implantation should be reserved for patients with significant electrocardiographic changes.

Calbindin D28K and parvalbumin gene expression in rat embryonic ventral forebrain grafts.

The present study characterizes expression of calbindin D28K (CB-D28K) and parvalbumin (PV) in ventral forebrain (VFB) grafts placed in the neocortex of adult rats bearing quisqualic acid lesions to the nucleus basalis magnocellularis. Three to nine months after transplantation surgery, rats were killed for in situ hybridization with probes to CB-D28K or PV and for immunohistochemistry with antibodies to CB-D28K or PV. In addition, an antibody to choline acetyltransferase (ChAT) was used to characterize the cholinergic component in the graft and an antibody to tyrosine hydroxylase (TH) to explore catecholaminergic innervation of the graft. Quantitative analysis of CB-D28K and PV messenger ribonucleic acid (mRNA) was based on counts of silver grains generated by emulsion autoradiography. Cells expressing CB-D28K mRNA were significantly larger than such cells in the adult VFB and the mean number of silver grains per cell was significantly greater than to such cells in the adult VFB. The level of CB-D28K mRNA expression as calculated by ratio of silver grains per unit area was also significantly increased. Quantification of PV mRNA showed no significant differences between the cells in the graft and in the adult VFB. In order to begin to interpret these findings, a comparison was made with such cells in the VFB of developing rats. Brain sections were sampled from embryonic day 17 and postnatal days 1, 5, 12, 19 and adult (6-12 months of age). Cells expressing CB-D28K mRNA were detected in ventral forebrain from postnatal day 5 and cells expressing PV mRNA were detected in ventral forebrain from postnatal day 19. In the course of normal development of the ventral forebrain, no CB-D28K cells were found that were as large or expressed such high levels of CB-D28K mRNA as observed in the grafts. We conclude that changes in grafted cells expressing CB-D28K do not reflect an arrest of developmental processes. TH immunohistochemistry revealed lack of catecholaminergic innervation of the graft, whereas adult mediolateral septal cells that express CB-D28K receive such innervation in addition to other neurotransmitter inputs. Imbalance in neurotransmitter inputs to grafted cells expressing CB-D28K is discussed as a possible factor in their increased size and gene expression.

Lossy (15:1) JPEG compression of digital coronary angiograms does not limit detection of subtle morphological features.

BACKGROUND: Development of the "all-digital" cardiac catheterization laboratory has been slowed by substantial computer archival and transfer requirements. Lossy data compression reduces this burden but creates irreversible changes in images, potentially impairing detection of clinically important angiographic features. METHODS AND RESULTS: Fifty image sequences from 31 interventional procedures were viewed both in the original (uncompressed) state and after 15:1 lossy Joint Photographic Expert's Group (JPEG) compression. Experienced angiographers identified dissections, suspected thrombi, and coronary stents, and their results were compared with those from a consensus panel that served as a "gold standard." The panel and the individual observers reviewed the same image sequences 4 months after the first session to determine intraobserver variability. Intraobserver agreement for original images was not significantly different from that for compressed images (89.8% versus 89.5% for 600 pairs of observations in each group). Agreement of individual observers with the consensus panel was not significantly different for original images from that for compressed images (87.6% versus 87.3%; CIs for the difference, -4.0%, 4.0%). Subgroup analysis for each observer and for each detection task (dissection, suspected thrombus, and stent) revealed no significant difference in agreement. CONCLUSIONS: The identification of dissections, thrombi, and coronary stents is not substantially impaired by the application of 15:1 lossy JPEG compression to digital coronary angiograms. These data suggest that digital angiographic images compressed in this manner are acceptable for clinical decision-making.

Role of right ventricular and pulmonary functional abnormalities in limiting exercise capacity in adults with congenital heart disease.

This study evaluates right ventricular (RV) and pulmonary function during exercise in adults with congenital heart disease (CHD). Thirty-one patients with CHD involving the right side of the heart underwent symptom-limited bicycle exercise testing with simultaneous expired gas analysis and measurement of RV ejection fraction (EF). Twenty-one age-matched normal controls underwent the identical exercise protocol. Maximal oxygen consumption was lower in the CHD than in normal controls (19.5 +/- 6.4 vs 30.5 +/- 0.8 ml/kg/min, p = 0.0001 patients vs controls). Both heart rate (156 +/- 25 vs 171 +/- 13 beats/min, p = 0.01) and oxygen pulse (9.3 +/- 3.7 vs 12.3 +/- 3.7 ml/beat, p = 0.01), an indirect measure of stroke volume, were found to be lower in the CHD group at peak exercise. Pulmonary dysfunction was evidenced in the CHD group by decreased forced expiratory volume, forced vital capacity and maximum voluntary ventilation, and by a higher ventilation/expired carbon dioxide ratio at peak exercise (37.2 +/- 6.9 vs 33.0 +/- 5.4, p = 0.02), suggesting an increase in dead space ventilation. Maximal oxygen consumption was lower in patients whose RVEF decreased with exercise (17.6 +/- 5.4 vs 22.8 +/- 6.4 ml/kg/min, p = 0.03 "decrease RVEF" group vs "increase RVEF" group). Maximal oxygen consumption correlated with the change in RVEF only in the group whose RVEF decreased with exercise (r = 0.5, p = 0.03). In the group that had increased RVEF with exercise, maximal oxygen consumption correlated with forced expiratory volume (r = 0.7, p = 0.02). Thus, adults with CHD have a reduced functional capacity compared with normal controls. This phenomenon appears to be associated with both RV and pulmonary abnormalities.

Effect of inadequate cardiac output reserve on exercise tolerance in patients with moderate mitral stenosis.

Twenty-nine patients with moderate mitral stenosis and 29 age-matched normal controls underwent symptom-limited upright bicycle exercise testing with simultaneous hemodynamic monitoring. Exercise tolerance in the mitral stenosis group was found to be limited by inadequate cardiac output reserve and not by resting mitral valve area or exercise pulmonary capillary wedge pressure.

Nitric oxide synthase in ventral forebrain grafts and in early ventral forebrain development.

Embryonic ventral forebrain (VFB) grafts to cortex contain neurons that synthesize acetylcholine and partially ameliorate behavioral deficits caused by excitotoxic damage to the nucleus basalis magnocelullaris in rats. An additional neurotransmitter, nitric oxide (NO), is synthesized by a subset of cholinergic neurons in rat ventral forebrain. If this neurotransmitter is expressed also by grafted cholinergic neurons (which include the embryonic medial septum and diagonal band), its functional contribution should be considered. Six to twelve months after transplantation of embryonic VFB tissue rats were sacrificed. Brain tissue was processed either for in situ hybridization of nNOS and neuropeptide Y (NPY) or for immunohistochemistry of choline acetyltransferase (ChAT) and neuronal nitric oxide synthase (nNOS). Quantification of messenger ribonucleic acid (mRNA) for nNOS was performed with radioactively labeled probes (silver grains were counted) and a preliminary comparison was made of graft sections to sections of the ventral forebrain of developing rats. Plots of silver grain counts against cell size revealed similar patterns in the grafts and in the ventral forebrain of developing rats. The rates of expression of mRNA for nNOS in the grafts were intermediate between those of the ventral forebrain of postnatal day 19 and those of postnatal day 12. Double immunohistochemical labeling revealed that 45.87 + 8.26% of cells expressing ChAT also expressed nNOS in the grafts, significantly higher than 33.16 + 3.9% which was the rate of co-expression observed in the adult ventral forebrain. This study suggests that possible contribution of NO to graft-associated modulation of behavior should be examined.

Esophageal varices in association with unilateral pulmonary vein atresia.

A 27-yr-old woman with recurrent episodes of hemoptysis (and hematemesis due to esophageal varices) was found to have unilateral pulmonary vein atresia. Reversed flow in the left pulmonary artery, a finding highly suggestive of this rare congenital anomaly, was demonstrated during cardiac catheterization. The definitive diagnosis was afforded by pulmonary wedge angiography, which not only demonstrated the atretic leftsided pulmonary veins, but also revealed a serpiginous system of systemic collateral veins to be the cause of her esophageal varices. Pneumonectomy of the nonfunctioning hypoplastic lung, the most successful approach to this disorder, may be complicated by severe bleeding of the numerous systemic arterial collateral vessels transected during surgical mobilization of the affected lung. Percutaneously delivered vascular occlusion coils were used in this case to occlude the large systemic to pulmonary artery collateral arteries immediately prior to pneumonectomy. Intra- and perioperative bleeding was minimal, and the patient has done well without further episodes of hemoptysis or hematemesis. Percutaneous occlusion of systemic to pulmonary collaterals may prove useful as a preoperative step for other congenital heart disease patients requiring lung or heart/lung transplantation.

Compression of digital coronary angiograms does not affect visual or quantitative assessment of coronary artery stenosis severity.

Digital coronary angiographic techniques are now widely used in many cardiac catheterization laboratories. However, the full potential of digital imaging technology remains to be achieved because of its enormous storage and exchange requirements. Compression of digital imaging data allows a reduction in the volume of data so that storage and transmission are more efficient and cost-effective. Three angiographers reviewed the original and compressed formats of 96 coronary angiographic sequences in a blinded fashion to assess coronary lesion severity. Compression was achieved using the Joint Photographic Experts Group (JPEG) standard, which resulted in a compression ratio of approximately 15:1. The original format was reviewed in a blinded fashion a second time to assess for intraobserver variability of similar formats. Lesion severity was graded in quartiles. Coronary stenosis >50% was considered "significant." In parallel, the reproducibility of quantitative coronary angiographic (QCA) measurements of coronary artery dimensions was also evaluated. For the visual assessment of lesion severity in the compressed versus original formats, kappa=0.52, suggesting moderate agreement. When lesions were assessed as significant versus "insignificant," however, kappa=0.88, suggesting excellent agreement. In the 2 separate readings of the original data formats, kappa=0.44 for assessment of lesion severity by quartiles and kappa=0.72 for lesions assessed as significant versus insignificant. Analysis of the compressed versus original data sets using QCA resulted in an excellent correlation for the measurement of lesion severity (r=0.99). The correlation was equally strong when the original format was analyzed sequentially (r=0.98). Lossy JPEG (15:1) compression is a valid means for reducing storage and exchange requirements of coronary angiographic data. The variability in assessing lesion severity between the original and compressed formats is comparable to the reported variability in visual assessment of lesion severity in sequential analysis of cine film.

Deficits in memory and hippocampal long-term potentiation in mice with reduced calbindin D28K expression.

The influx of calcium into the postsynaptic neuron is likely to be an important event in memory formation. Among the mechanisms that nerve cells may use to alter the time course or size of a spike of intracellular calcium are cytosolic calcium binding or "buffering" proteins. To consider the role in memory formation of one of these proteins, calbindin D28K, which is abundant in many neurons, including the CA1 pyramidal cells of the hippocampus, transgenic mice deficient in calbindin D28K have been created. These mice show selective impairments in spatial learning paradigms and fail to maintain long-term potentiation. These results suggest a role for calbindin D28K protein in temporally extending a neuronal calcium signal, allowing the activation of calcium-dependent intracellular signaling pathways underlying memory function.

Total pulmonary artery occlusion immediately after lung transplantation: successful revascularization with intravascular stents.

Occlusion of the pulmonary artery anastomosis is an uncommon early complication of lung transplantation which necessitates emergency surgical revision. Percutaneous correction of such a complication has not been previously reported. We report a patient who had total pulmonary artery occlusion less than 24 hours after right single lung transplantation with successful revascularization achieved by means of percutaneously delivered endovascular stents.

Left ventricular performance improves late after aortic valve replacement in patients with aortic stenosis and reduced ejection fraction.

EF in patients with aortic stenosis and reduced EF who underwent aortic valve replacement did not improve by 1 week postoperatively despite rectification of afterload mismatch. By 6 months, however, EF significantly improved without any further change in ventricular loading conditions. This implies that the benefit from aortic valve replacement (when measured by LV ejection performance) may not be evident until late postoperatively.

Apolipoprotein D gene polymorphism: a new genetic marker for type 2 diabetic subjects in Nauru and south India.

Type 2 diabetes is characterized by abnormalities in both glucose and lipoprotein metabolism and genes involved in lipid metabolism are legitimate candidates for involvement in Type 2 diabetes. We have previously reported an association in Nauruans between a Taq 1 polymorphism of the apolipoprotein D gene (apo D) and Type 2 diabetes. In this study these findings were investigated further in the Nauruan population as well as two other ethnic groups. In South Indian subjects, there was a significant difference in genotype distribution of apo D genotypes between diabetic subjects (n = 110) and controls (n = 88; p = 0.004) which was similar to that previously found in the Nauruan subjects. No such association was seen in elderly Finnish subjects (diabetic n = 69; impaired glucose tolerance n = 26 and normal glucose tolerance n = 31). Linkage between the apo D polymorphism and diabetes in 12 Nauruan families was only excluded under a highly penetrant dominant model and was unlikely under other single gene models. Since the beta cell glucose transporter gene (Glut 2) is found in a similar chromosomal location to apo D, South Indian subjects (diabetic n = 95 and controls n = 56) were typed at this locus. No association between diabetes and the Glut 2 Taq I polymorphism was found in the South Indian subjects. Furthermore, there was no evidence of linkage disequilibrium between the apo D and Glut 2 genes. In conclusion, apo D might act as a modifying gene for Type 2 diabetes in some ethnic groups.

Exploratory celiotomy for suspected urinary tract disruption in neonatal foals: a review of 18 cases.

The medical records of 18 neonatal foals, in which exploratory celiotomies were performed for suspected urinary tract lesions, were reviewed. Despite clinical signs and laboratory values indicative of disruption of the urinary tract, three foals did not have a site of urinary tract leakage at surgery. Eight foals had ruptured bladders and seven foals had urachal lesions. Ultrasonography was used as a pre-operative diagnostic procedure in eight foals to evaluate the presence of free peritoneal fluid and urinary tract integrity. Nine foals were alive six months after discharge. Seven of the nine non-surviving foals died or were destroyed because of fungal or bacterial infections.

Exploratory celiotomy for gastrointestinal disease in neonatal foals: a review of 20 cases.

The medical records of 20 neonatal foals in which exploratory celiotomies were performed for gastrointestinal disease were reviewed. In all 20 foals, persistent pain and/or progressive abdominal distension were the primary clinical findings influencing the decision to operate. However, ancilliary laboratory data were important to the proper medical management of these foals during anaesthesia and following surgery. Surgical diagnoses of the 20 foals included ileus (nine foals; 45 per cent), small colon obstruction (five foals; 25 per cent), large colon displacement (three foals; 15 per cent), small intestinal displacement (two foals; 10 per cent), and perforated gastric ulcer (one foal; 5 per cent). Seventeen foals were recovered from anaesthesia, 13 of these were discharged from the hospital, seven were alive six months or more following discharge. Sepsis was the cause of death in six of the 10 foals that died following recovery.

Penicillin G susceptibility of "rural" staphylococcus aureus.

A total of 9,955 isolates of Staphylococcus aureus (SA) from an urban hospital and clinic and from rural office settings obtained during 1971 through 1978 were compared for susceptibility to penicillin G. Clear downward trends in susceptibility of SA were noted for all urban settings during the 1970s. In 1978, penicillin G susceptibility of SA in an urban community practice (16.3 percent) was actually less than that in the urban hospitals in the same city (17.1 percent). Penicillin G susceptibility of SA from two rural sources was likewise very low (20.9 percent and 28.7 percent). Therefore, Staphylococcus aureus must be considered resistant to penicillin G until proven otherwise, irrespective of the source of the isolate.