Effective intracellular delivery of oligonucleotides in order to make sense of antisense.

For more than two decades, antisense oligonucleotides (ODNs) have been used to modulate gene expression for the purpose of applications in cell biology and for development of novel sophisticated medical therapeutics. Conceptually, the antisense approach represents an elegant strategy, involving the targeting to and association of an ODN sequence with a specific mRNA via base-pairing, resulting in an impairment of functional and/or harmful protein expression in normal and diseased cells/tissue, respectively. Apart from ODN stability, its efficiency very much depends on intracellular delivery and release/access to the target side, issues that are still relatively poorly understood. Since free ODNs enter cells relatively poorly, appropriate carriers, often composed of polymers and cationic lipids, have been developed. Such carriers allow efficient delivery of ODNs into cells in vitro, and the mechanisms of delivery, both in terms of biophysical requirements for the carrier and cell biological features of uptake, are gradually becoming apparent. To become effective, ODNs require delivery into the nucleus, which necessitates release of internalized ODNs from endosomal compartments, an event that seems to depend on the nature of the delivery vehicle and distinct structural shape changes. Interestingly, evidence is accumulating which suggests that by modulating the surface properties of the carrier, the kinetics of such changes can be controlled, thus providing possibilities for programmable release of the carrier contents. Here, consideration will also be given to antisense design and chemistry, and the challenge of extra- and intracellular barriers to be overcome in the delivery process.     

Can Electrocardiographic Criteria Predict Adverse Cardiac Events and Positive Cardiac Markers?

OBJECTIVES: To determine electrocardiogram (ECG) predictors of positive cardiac markers and short-term adverse cardiac events in an undifferentiated chest pain population presenting to emergency departments (EDs). The authors hypothesized that specific ECG findings, other than those previously identified in higher-risk populations, would be predictive of cardiac outcomes and positive cardiac markers. METHODS: This study used data from a prospectively collected, retrospectively analyzed Internet-based data registry of undifferentiated chest pain patients (i*trACS). Logistic regression modeling was performed to determine the ECG findings that were predictive of 1) positive cardiac markers and 2) short-term adverse cardiac events. RESULTS: ST-segment elevation (STE), ST-segment depression (STD), pathological Q-waves (PQW), and T-wave inversion were associated with increased odds of percutaneous coronary intervention or catheterization, myocardial infarction, or coronary artery bypass grafting. The odds of creatine kinase-MB (CK-MB) measuring positive were increased if STE, STD, or PQW were present [odds ratio (OR) 2.495, 2.582, and 1.295, respectively]. A right bundle branch block tended to decrease the odds of CK-MB measuring positive (OR 0.658). A similar pattern of results was observed for troponin I (OR 3.608 for STE, 3.72 for STD, 1.538 for PQW). Troponin T showed an increased odds of measuring positive if any of STE, STD, left bundle branch block, or T-wave inversion were evident (OR 2.313, 2.816, 1.80, and 1.449, respectively). CONCLUSIONS: Initial ECG criteria can be used to predict short-term cardiac outcomes and positive cardiac markers. These findings can be important aids in the risk-stratification and aggressive treatment regimens of chest pain patients presenting to EDs.     

Specific MR imaging of human lymphocytes by monoclonal antibody-guided dextran-magnetite particles.

Human lymphocytes were labeled with biotinylated anti-lymphocyte-directed monoclonal antibodies, to which streptavidin and subsequently biotinylated dextran-magnetite particles were coupled. This labeling resulted in a strong and selective negative contrast enhancement of lymphocyte suspensions at 2.0 T, caused predominantly by the specific increase of R2 with a small but significant specific increase of R1. The R1 was found to decrease with increasing field strength. The immunolabeling procedure described here may be used for the selective signal depletion of target cells in MR imaging.     

Detection of local recurrence of soft-tissue sarcoma with positron emission tomography using [18F]fluorodeoxyglucose

Background: It is often difficult to detect a local recurrence of soft-tissue sarcomas due to disturbance of the normal anatomy by previous surgery and radiotherapy. The aim of this study was to assess the value of positron emission tomography (PET) with [¹⁸F]fluoro-2-deoxy-d-glucose (FDG) for detecting local recurrences. Methods: In the period 1992–1995, 17 patients with proven or suspected local recurrence of soft-tissue sarcoma were examined using FDG-PET. Fifteen of these patients were ultimately proven to have a recurrence. Results: Recurrence was visualized in 14 patients (93%). Small tumors (maximum diameter 0.5 cm) were as easily visible as large lesions (maximum diameter 20 cm). In one patient the PET scan was positive, but the recurrence could not be proven histologically. Recurrence was proven 1 year later. A recurrent low-grade liposarcoma was not visualized. The two patients with benign lesions had a negative PET scan. The mean glucose metabolic rate was calculated to be 13.2 μmol/100 g/min (range 1.9–28.4). A correlation was found between the histological malignancy grade and the metabolic rate (p<0.05; Kruskal-Wallis). Conclusion: PET with FDG is a useful addition to the diagnostic armamentarium for detecting local recurrence of soft-tissue sarcomas and provides an indication of the malignancy grade of the recurrent lesion. Presented at the 47th Annual Meeting of The Society for Surgical Oncology, Houston, Texas, March 17–20, 1994.     

Respiratory atopic disease, Ascaris-immunoglobulin E and tuberculin testing in urban South African children

BACKGROUND: Epidemiological relation of intestinal helminth infection and atopic disease, both associated with a T-helper (Th) 2 immune response, is controversial, as it has been reported that helminth infection may either suppress or pre-dispose to atopic disease. This relation has not been tested in an area with a high burden of Mycobacterium tuberculosis (MTB) infection, a known Th1-stimulating infection. OBJECTIVE: To study the association of intestinal helminth infection and atopic disease in a community where helminth infection is endemic and MTB infection is high. METHODS: Three-hundred and fifty-nine randomly selected children aged 6-14 years from a poor urban suburb were tested with allergy questionnaire, skin prick test (SPT) to common aeroallergens, Ascaris-specific IgE (Ascaris-sIgE), fecal examination for pathogenic intestinal helminths and tuberculin skin testing (TST). Histamine bronchoprovocation was tested in the group of children aged 10 years and older. RESULTS: were corrected for demographic variables, socioeconomic status, parental allergy, environmental tobacco smoke (ETS) exposure in the household, recent anthelminthic treatment and for clustering in the sampling unit. Results Ascaris-sIgE was elevated in 48% of children, Ascaris eggs were found in 15% and TST was positive in 53%. Children with elevated Ascaris-sIgE had significantly increased risk of positive SPT to aeroallergens, particularly house dust mite, atopic asthma (ever and recent), atopic rhinitis (ever and recent) and increased atopy-related bronchial hyper-responsiveness. In children with negative TST (<10 mm), elevated Ascaris-sIgE was associated with significantly increased risk of atopic symptoms (adjusted odds ratio (OR(adj)) 6.5; 95% confidence interval (CI) 1.9-22.4), whereas in those with positive TST (>/=10 mm) this association disappeared (OR(adj) 0.96; 95% CI 0.4-2.8). CONCLUSIONS: These results suggest that immune response to Ascaris (Ascaris-sIgE) may be a risk factor of atopic disease in populations exposed to mild Ascaris infection and that MTB infection may be protective against this risk, probably by stimulation of anti-inflammatory networks.     

Nocturnal hypoglycaemia in Type 1 diabetic patients,assessed with continuous glucose monitoring: frequency,duration and associations

Aims We quantified the occurrence and duration of nocturnal hypoglycaemia in individuals with Type 1 diabetes treated with continuous subcutaneous insulin infusion (CSII) or multiple‐injection therapy (MIT) using a continuous subcutaneous glucose sensor. Methods A microdialysis sensor was worn at home by 24 patients on CSII (mean HbA_1c 7.8 ± 0.9%) and 33 patients on MIT (HbA_1c 8.7 ± 1.3%) for 48 h. Occurrence and duration of nocturnal hypoglycaemia were assessed and using multivariate regression analysis, the association between HbA_1c, diabetes duration, treatment type (CSII vs. MIT), fasting and bedtime blood glucose values, total daily insulin dose and mean nocturnal glucose concentrations, and hypoglycaemia occurrence and duration was investigated. Results Nocturnal hypoglycaemia ≤ 3.9 mmol/l occurred in 33.3% of both the CSII‐ (8/24) and MIT‐treated patients (11/33). Mean (± sd ; median, interquartile range) duration of hypoglycaemia ≤ 3.9 mmol/l was 78 (± 76; 57, 23–120) min per night for the CSII‐ and 98 (± 80; 81, 32–158) min per night for the MIT‐treated group. Multivariate regression analysis showed that bedtime glucose value had the strongest association with the occurrence (P = 0.026) and duration (P = 0.032) of nocturnal hypoglycaemia. Conclusions Microdialysis continuous glucose monitoring has enabled more precise quantification of nocturnal hypoglycaemia occurrence and duration in Type 1 diabetic patients. Occurrence and duration of nocturnal hypoglycaemia were mainly associated with bedtime glucose value.     

Effect of ambient oxygen concentration upon the acute toxicity of chlorophenols and heavy metals to the groundwater copepod Parastenocaris germanica (Crustacea).

Acute static toxicity experiments have been performed under normoxic and hypoxic conditions. The test animals used were adults of the groundwater-adapted copepod Parastenocaris germanica. The animals originated from a sandy, gravelly phreatic aquifer of the Meuse valley in The Netherlands. Toxicants applied were pentachlorophenol, 3,4-dichlorophenol, zinc, and cadmium. The results were statistically evaluated by a log-logistic model. LC50 and LC5 values with 95% confidence limits were calculated. Response models of application under both oxygen conditions were compared. The influence of ambient oxygen concentration on sensitivity appeared to be very small; only in the case of pentachlorophenol were hypoxic and normoxic models slightly but significantly different. Comparisons with the sensitivity of other organisms are made and the results are discussed against the specific physiological adaptations of the organisms to groundwater conditions.