Tumor necrosis factor-alpha and interleukin-1beta mediate endothelial permeability induced by lipopolysaccharide-stimulated whole blood

OBJECTIVE: To investigate the role of endotoxin-induced inflammatory mediators in blood on the permeability of endothelial monolayers. DESIGN: Whole blood of healthy volunteers was treated with bacterial lipopolysaccharide (Escherichia coli, B55:05), and the resultant plasma was added to human umbilical venular endothelial cells (HUVEC) cultured on semipermeable membrane inserts (Transwells). SETTING: University hospital laboratory. SUBJECTS: Whole blood of healthy volunteers. INTERVENTIONS: Donor plasma was treated with excess antibodies against either tumor necrosis factor-alpha, interleukin-1beta, or both, before the incubation on HUVEC. MEASUREMENTS AND MAIN RESULTS: The permeability of HUVEC monolayers to fluorescent-labeled albumin and dextran was measured over a 6-hr period, after removal of the stimulus. The production of tumor necrosis factor-alpha and interleukin-1beta in lipopolysaccharide-treated whole blood was determined by radioimmunoassay. Individually, lipopolysaccharide (10 microg/mL), tumor necrosis factor-alpha (10 ng/mL), and interleukin-1beta (50 ng/mL) all increased endothelial permeability by about 2.5-fold. A much larger increase could be achieved by preincubation of lipopolysaccharide (10 microg/mL) in whole blood: the resultant plasma induced a ten-fold increase of the permeability. The permeability response after preincubation of lipopolysaccharide in whole blood was time- and dose-dependent. Moreover, this treatment increased the sensitivity of endothelial monolayers to lipopolysaccharide by a factor of several thousand-fold: Whereas high doses of lipopolysaccharide were required for direct stimulation of the permeability, picomolar amounts of lipopolysaccharide in whole blood induced a similar increase. Significant amounts of tumor necrosis factor-alpha and interleukin-1beta were produced in blood at similar doses of lipopolysaccharide. The addition of antibodies against tumor necrosis factor-alpha or interleukin-1beta to plasma partially but significantly abrogated the permeability increase. However, a complete inhibition could be achieved by the simultaneous addition of anti-tumor necrosis factor-alpha and anti-interleukin-1beta to plasma. CONCLUSIONS: Although lipopolysaccharide is capable of directly inducing endothelial permeability, blood-borne tumor necrosis factor-alpha and interleukin-1beta mediate lipopolysaccharide-induced endothelial permeability at low endotoxin concentrations. These findings support the idea that multifactorial inhibition of inflammatory mediators may improve survival in septic patients.     

The release of gentamicin from acrylic bone cements in a simulated prosthesis-related interfacial gap

Gentamicin is added to polymethylmethacrylate bone cements in orthopedics as a measure against infection in total joint arthroplasties. Numerous studies have been published on gentamicin release from bone cements, but none have been able to estimate the local concentrations in the prosthesis-related interfacial gap, present after implantation. The aim of this study was to develop a method allowing determination of antibiotic release in such a gap. Two-hundred-micrometer-wide gaps with a volume of 6 microl and a surface area of 0.6 cm2 were created by inserting stainless-steel strips in gentamicin-loaded bone cement plugs prior to polymerization. After hardening, the gap surface was exposed to 6 microl or 10 ml of phosphate-buffered saline. Within 2 h, gentamicin concentrations in the gaps reached around 4000 microg/ml for 4 different CMW and Palamed cements and 2500 microg/ml for Palacos R. Concentrations measured in the larger volume were several hundred times lower than in the gaps. This simulated prosthesis-related interfacial gap model offers new insights in the clinical efficacy of antibiotic-loaded bone cements. It is demonstrated that concentrations up to 1000-fold the antibiotic resistance levels for most bacterial strains causing implant infection can be achieved in a realistic in vitro model.     

Coregulation of epidermal growth factor receptor/human epidermal growth factor receptor 2 (HER2) levels and locations: quantitative analysis of HER2 overexpression effects

Elevated expression of human epidermal growth factor receptor 2 (HER2) is known to alter cell signaling and behavioral responses implicated in tumor progression. However, multiple diverse mechanisms may be involved in these overall effects, including signaling by HER2 itself, modulation of signaling by epidermal growth factor receptor (EGFR), and modification of trafficking dynamics for both EGFR and HER2. Because these processes are so tightly interrelated, the net effect of HER2 overexpression is difficult to reliably attribute to any single particular mechanism. To take an important first step toward dissecting the effects of HER2 overexpression on cell responses in terms of the various specific underlying mechanisms, we have developed and validated a quantitative model of the relevant trafficking processes. We then use our model for successful prediction of EGFR and HER2 level and location changes attributable to HER2 overexpression in 184A1 human mammary epithelial cells expressing a series of HER2 levels by retroviral infection. Model predictions are based on our independent experimental measurement of key trafficking parameters for both EGFR and HER2. In terms of trafficking processes, HER2 overexpression reduces the EGFR internalization rate constant and increases the fraction of EGFR recycled. Consequently, our model successfully predicts that HER2 increases the overall level of activated EGFR by both enhancing its recycling and reducing its internalization, but it increases activated EGFR localization at the cell surface almost solely by its reduction of internalization. Furthermore, the model also successfully predicts the effects of monoclonal antibody 2C4, which interferes with HER2/EGFR heterodimerization, on EGFR and HER2 levels and compartmental locations. We anticipate that this model should ultimately be useful in parsing the relative contributions of direct effects of HER2 via signaling vis-a-vis indirect effects of HER2 via modification of EGFR signaling.     

Recognition memory of serially or simultaneously presented words or figures, of epilepsy patients with or without mesial temporal sclerosis

Previous studies, examining short-term recognition memory in patients with partial seizures as a consequence of mesial temporal sclerosis (MTS) have reported inconsistent findings. Dependent on the paradigms used for measuring recognition memory, some studies have demonstrated that the mesial temporal structures are not critically involved in short-term recognition memory. In addition, other studies found a lateralization effect that is consistent with the generally accepted association between left temporal lobe lesions and verbal memory deficits, and right temporal lobe lesions and non-verbal memory impairments. In the present study verbal and non-verbal recognition memory was tested in 41 patients with left or right temporal lobe epilepsy with MTS (22 left; 19 right) versus 44 patients with left or right temporal lobe epilepsy but without MTS (28 left; 16 right). Verbal and non-verbal recognition tasks were presented in both a serial and simultaneous condition to test a hypothesized local/global paradigm. Multiple analyses of variance (MANOVA) showed that unilateral MTS has no marked effects on verbal or non-verbal recognition memory of patients with seizures. An interaction effect between MTS and the lateralization of epileptic activity was found on memory performance: MTS only leads to deficits in recognition memory in patients with right-sided epileptiform activity. As hypothesized, patients with left temporal lobe abnormalities, have specific deficits with recognizing serially presented information.     

Vitamin K supplementation in cystic fibrosis

The relation between different doses of vitamin K supplementation, several bone markers, and PIVKA-II concentrations in cystic fibrosis (CF) patients compared to controls was evaluated. Results suggest that a increased vitamin K intake may have significant health benefits for children with CF.     

Breast carcinomas of limited extent: frequency, radiologic-pathologic characteristics, and surgical margin requirements

BACKGROUND: Clinical trials established the value of breast-conserving treatment (BCT) including the macroscopic removal of the tumor followed by local radiation therapy (RT) for Stage I and II invasive carcinomas. The occurrence of local tumor recurrence is related to the extent and multifocality of the tumor. Various studies aim to identify those tumors that could be proper candidates for conventional BCT. Furthermore, recent studies have focused on the identification of tumors that may be treated by breast-conserving surgery alone without RT. Small, localized tumors theoretically should be the potential candidates for this type of treatment. The mammographic and pathologic criteria for the identification of tumors with limited extent are not yet established; furthermore, the optimal extent of the surgical excision and the method for margin examination are controversial. METHODS: Surgical breast-conserving procedures were simulated in a review of 135 mastectomy specimens of patients treated for an invasive carcinoma (> or = 4 cm in size, all pathologic types except invasive lobular carcinoma) who were theoretically eligible for conservative treatment. Tumor spread including possible multifocality and multicentricity was studied by the technique of correlated specimen radiography and pathology. Breast carcinoma of limited extent (BCLE), the proper tumor profile for BCT, was defined as having no invasive carcinoma, ductal carcinoma in situ, and lymphatic emboli foci beyond 1 cm from the edge of the dominant mass. RESULTS: Fifty-three percent of the patients in this series had a BCLE. No statistically significant relation was found between BCLE and patient age, pathologic size, type and grade of the tumor, lymph node status, mode of detection, and mammographic aspect of the index tumor. Based on mammography, the absence of calcification or tumor density beyond the edge of index tumor appears to be the best predictor for BCLE (P < 0.0001). A 1-cm microscopically tumor free margin as the outer rim of a macroscopic surgical margin of 2 cm gives the best positive predictive value based on pathology (P < 0.0001). By applying the above conditions, 72 of the 135 cancers were identified as being potential BCLE cases in this series. However, whereas 64 of these 72 tumors (89%) were correctly identified as being true BCLE, 8 (11%) were erroneously identified as such (non-BCLE cases), having "residual" tumor foci beyond 2 cm from the edge of the dominant tumor. CONCLUSIONS: We conclude, that approximately 50% of invasive ductal carcinomas may have limited extent. The accuracy of identifying this group of cancers, the proper candidates for BCT, by applying state-of-the-art mammography and pathology may be as high as 90%. A subset of these tumors might represent the potential candidates for treatment with surgery alone without RT. As a result, the routine application of BCT complemented by RT would have led to the overtreatment of 89% of the patients with a BCLE in this series; conversely, 11% of the tumors may have recurred without the use of RT. Considering that these conclusions are based on a theoretic morphologic model, further clinical studies with facilities for high quality team approach in diagnosis and therapy are needed to evaluate the impact of BCLE on BCT strategies. The results of this study should not justify the withholding of RT outside the context of clinical trials.     

Parents' responses to disclosure of genetic test results of their children

The psychological reactions of 22 parental couples and 3 single parents were investigated after disclosure of genetic test results of their children. The children were tested for the early-onset, monogenetic cancer disorder multiple endocrine neoplasia type 2. Participants came from 13 different families and were aged between 28 and 47 years. Parents who were informed that their child was a gene carrier reacted with resignation, showed moderate to high levels of test-related and general anxiety, but few psychological complaints. Daily activities were disturbed in 43% of the parents with carrier-children. There was little disruption of the parents' future perspective, apart from some socioeconomic disadvantages and increased parental concern for the carrier-children. Most parents with carrier-children showed restraint with respect to short-term prophylactic treatment. Parents with favorable test results showed significantly less anxiety and no disturbance in their daily activities. They did not, however, seem to be reassured by the DNA test result. These parents questioned the reliability of the DNA test, wanted confirmation of the test results, and were eager to continue screening of their noncarrier children. Parents, especially those with a lower level of education and/or a pessimistic view of the future, were distressed by unfavorable test results. Additional counseling is advised to prevent parents of carrier-children worrying unnecessarily, or parents with children in whom the disease gene was not found being not reassured. Am. J. Med. Genet. 94:316-323, 2000.     

Facilitation of memory by contextual cues in patients with diencephalic or medial temporal lobe dysfunction

Item-context binding is crucial for successful episodic memory formation, and binding deficits have been suggested to underlie episodic-memory deficits. Here, our research investigated the facilitation of cued recall and recognition memory by contextual cues in 20 patients with Korsakoff's amnesia, 20 unilateral medial-temporal lobectomy (MTL) patients and 36 healthy controls. In a computerized task participants had to learn 40 nouns that were randomly combined with a photograph of an everyday scene. Korsakoff patients showed a general memory deficit in both the cued recall and the recognition condition. A less severe memory impairment was found in the patients with medial-temporal lobectomy. Contextual cues facilitated cued recall to an equal extent in unilateral temporal lobectomy patients and healthy controls. However, no facilitation was observed in Korsakoff patients, suggesting an impairment in item-context binding during cued recall tasks. In contrast to the presumed exclusive dependency of recognition memory on item information, all groups equally profited from the contextual cues in recognition tasks. Our findings show that unilateral lesions as with MTL result in normal binding of context and item information, while bilateral dysfunction of the hippocampal-diencephalic system results in impaired context and item binding.