Crystal structure of TNF-alpha mutant R31D with greater affinity for receptor R1 compared with R2

Crystal structures have been determined of recombinant human tumor necrosis factor-alpha (TNF-alpha) and its R31D mutant that preferentially binds to TNF receptor R1 with more than seven times the relative affinity of binding to receptor R2. Crystals of the wild-type TNF were of space group P4(1)2(1)2 and had unit cell dimensions of a = b = 94.7 and c = 117.4 A. Refinement of the structure gave an R-factor of 22.3% at 2.5 A resolution. The crystals of TNF R31D mutant diffracted to 2.3 A resolution, and were of identical space group to the wild type with unit cell dimensions of a = b = 95.4 and c = 116.2 A, and the structure was refined to an R-factor of 21.8%. The trimer structures of the wild-type and mutant TNF were similar with a root mean square (r.m.s.) deviation of 0.56 A for Calpha atoms; however, the subunits within each trimer were more variable with an average r.m.s. deviation of 1.00 A on Calpha atoms for pairwise comparison of subunits. Model complexes of TNF with receptors R1 and R2 have been used to predict TNF-receptor interactions. Arg31 in all three subunits of wild-type TNF is predicted to form an ionic interaction with the equivalent glutamic acid in both receptors R1 and R2. Asp31 of the TNF R31D mutant is predicted to interact differently with the two receptors. The side chain of Asp31 in two subunits of the TNF mutant is predicted to form hydrogen bond interactions with Ser59 or Cys70 of R1, while it has no predicted interactions with R2. The loss of three strong ionic interactions of Arg31 and the electrostatic repulsion of Asp31 with Glu in the receptors is consistent with the reduced binding of the R31D mutant to both receptors relative to wild-type TNF. The replacement of these ionic interactions by two weaker hydrogen bond interactions between Asp31 of the R31D mutant and R1, compared with no interactions with R2, is in agreement with the observed preferential binding of the R31D mutant to R1 over R2. Analysis of the structure and function of receptor-discriminating mutants of TNF will help understand the biological role of TNF and facilitate its use as an antitumor agent.      

Protecting Business Secrets in an EDI/CALS Environment

ABSTRACT

Communicating with confidential data requires special attention in a mobile agents environment, especially when the other hosts must be prevented from eavesdropping on the communication. We propose a communication model for secured communication between the agents belonging to publishers and consumers data. Confidentiality is ensured using our on-the-fly encryption-decryption sequence using ElGamal system to directly convert the message or plaintext into one that is encrypted directly with the public key of consumer. The scheme ensures that the data possessed by the agents is secured at all times when it is executing at any untrusted host. Our minimal implementation of the model with Aglets agent platform gives the first faithful picture of the happenings in the model. Finally, we also explain how the homomorphic property of ElGamal scheme can be integrated with our model for a Web-based application such as voting involving multiple agents.     

Time is of the essence: a conjecture that hemispheric specialization arises from interhemispheric conduction delay

Tomasch (1954) and Aboitiz et al. (1992) found the majority of the fibers of the human corpus callosum are under 1 micron in diameter. Electron microscopic studies of Swadlow et al. (1980) and the detailed study of LaMantia and Rakic (1990a) on macaques show the average size of the myelinated callosal axons also to be less than 1 micron. In man, the average-sized myelinated fiber interconnecting the temporal lobes would have a one-way, interhemispheric delay of over 25 msec. Thus, finely detailed, time-critical neuronal computations (i.e., tasks that strain the capacity of the callosum and hence could not be handled by just the larger fibers) would be performed more quickly via shorter and faster intrahemispheric circuits. While one transit across the commissural system might yield tolerable delays, multiple passes as in a system involving "setting" would seem prohibitively slow. We suggest that these temporal limits will be avoided if the neural apparatus necessary to perform each high-resolution, time-critical task is gathered in one hemisphere. If the, presumably overlapping, neural assemblies needed to handle overlapping tasks are clustered together, this would lead to hemispheric specialization. The prediction follows that the large brains of mammals such as elephants and cetaceans will also manifest a high degree of hemispheric specialization.     

Correction of congenital diaphragmatic hernia in utero IX: fetuses with poor prognosis (liver herniation and low lung-to-head ratio) can be saved by fetoscopic temporary tracheal occlusion

BACKGROUND/PURPOSE: Fetuses with congenital diaphragmatic hernia (CDH) who have a "poor prognosis" with postnatal treatment now can be identified on the basis of liver herniation, early diagnosis (before 25 weeks' gestation) and a low lung-to-head ratio (LHR). Because complete in utero repair proved unsuccessful for this group, the strategy of temporary tracheal occlusion was developed to gradually enlarge the hypoplastic fetal lung. The purpose of this study is to compare the outcome of patients in the poor-prognosis group treated by one of three methods: (1) standard postnatal care, (2) fetal tracheal occlusion via open hysterotomy, and (3) the recently developed video-fetoscopic (Fetendo) technique of tracheal occlusion without hysterotomy. METHODS: In the past 3 years, 34 of 86 fetuses with an isolated left CDH met criteria for the poor-prognosis group. Thirteen families chose postnatal treatment at an extracorporeal membrane oxygenation (ECMO) center, 13 underwent open fetal tracheal occlusion, and eight underwent fetoscopic tracheal occlusion. RESULTS: The survival rate was 38% in the group treated by standard postnatal therapy, 15% in the open tracheal occlusion group, and 75% in the Fetendo group. There were less postoperative pulmonary complications noted in mothers who underwent the Fetendo procedure versus the open tracheal occlusion. All but one Fetendo clip patient had a striking physiological response demonstrated by sonographic enlargement of the small left lung that was documented postnatally by plain radiographs and its subjective appearance during repair of the CDH. In contrast, only 5 of the 13 open tracheal occlusion patients demonstrated lung growth. CONCLUSION: Fetuses with a left CDH who have liver herniation and a low LHR are at high risk of neonatal demise and appear to benefit from temporary tracheal occlusion when performed fetoscopically, but not when performed by open fetal surgery.     

A systems model for low vision rehabilitation. II. Measurement of vision disabilities

BACKGROUND: This study presents and tests a method of measuring vision disabilities. Based on the model presented in the first paper of this series, cognitive and motor activities ("tasks") are organized in a hierarchy that identifies the tasks' behavioral goals (purposes) and the social objectives the goals serve. Two latent variables important to the definition of vision disability are identified: "value of living independently" and "visual ability for independent living." The basic principles of the Rasch measurement model are reviewed, and Rasch models are used to measure the two variables that define vision disability. The need for rehabilitation to meet each goal is represented by "rehabilitative demand," a mathematical function of the measurements made of the value and difficulty of achieving each goal independently. METHODS: Over 400 patients with low vision rated both the importance and difficulty of independently achieving each of 24 goals. Rasch analysis was used to derive interval measures of the social value of each goal, the value that individual patients placed on each goal relative to the derived social definition of independence, the visual ability required to achieve each goal without help, and the visual ability of individual patients to live independently. A rehabilitative demand function was ascertained from the judgments of 17 AAO Diplomates in Low Vision through triadic comparisons and multidimensional scaling. RESULTS: Self-care had the greatest "social value" for independence; performing music had the least. Recreational reading required the greatest "visual ability" to accomplish independently; self-care required the least. Rehabilitative demand was linear with value and nonmonotonic with difficulty. CONCLUSIONS: Rehabilitative demand, an algorithm for defining vision disability, incorporates interval measures of visual ability and the value of independent living estimated from patient-based assessments, a social scale of the value of activities relative to independent living, and a consensus opinion of low vision experts on the prioritization of the need for rehabilitation.     

Comparison of CT and MR imaging in staging of neck metastases

OBJECTIVE: To determine whether adaptive cytoprotection exists in human intestinal cells under in vitro conditions and what role, if any, endogenous prostaglandins or calcium may play in mediating this protective response. SUMMARY BACKGROUND DATA: Adaptive cytoprotection can be defined as that process whereby the administration of a low concentration of a damaging agent, termed a "mild irritant," which by itself is not injurious, can attenuate gastrointestinal mucosal injury subsequently induced by the application of higher concentrations of the same or other necrotizing agents. Despite substantial investigation, the mediator or mediators of adaptive cytoprotection remain poorly understood. METHODS: Postconfluent Caco-2 cells were used in all experiments. Cellular death was quantitated using a dual-component fluorescent assay. Changes in intracellular calcium concentration were quantitated by measuring fluorescent signal changes of the single wavelength calcium indicator (Fluo-3). Finally, prostaglandin E2 release into the media was quantitated by radioimmunoassay. RESULTS: Pretreatment of Caco-2 cells with low concentrations of ethanol (mild irritant) significantly attenuated injury induced by higher damaging concentrations of ethanol. The protection conferred by the mild irritant was directly dependent on both the concentration of the irritant used and the duration of exposure and was abrogated when cells were pretreated with an endogenous prostaglandin inhibitor (indomethacin) or if the mild irritant was administered in calcium-free media. Cells exposed to ethanol had a significant and concentration-dependent increase in intracellular calcium concentration, an effect that was highly related to cellular injury. Pretreatment with a mild irritant significantly decreased intracellular calcium increases induced by not only ethanol but also by a calcium ionophore (A23187). Cells treated with low concentrations of ethanol demonstrated no significant elevation in prostaglandin E2 release. CONCLUSIONS: Adaptive cytoprotection induced by ethanol exists in human colonocytes under in vitro conditions independent of mucosal blood flow, neural innervation, or circulating humoral factors. The authors' data suggest that this response does not require endogenous prostaglandin synthesis but may involve processes whereby intracellular calcium accumulation is prevented.     

Plasma-derived proteins in airway defence, disease and repair of epithelial injury

One significant characteristic of the airway mucosa in vivo, that cannot easily be mimicked in vitro, is its microcirculation, which generates a highly dynamic, biologically active milieu of plasma-derived molecules that may pass to the airway lumen in vivo. New data on the mechanisms of airway mucosal exudation indicate that the protein systems of circulating plasma may contribute significantly to the biology and immunology of the lamina propria, its surface epithelium and the luminal surface, not only in injured airways, but also in airways that are activated but display no sign of oedema, epithelial disruption, or increased absorption capacity. We suggest that present knowledge of the mechanisms of plasma exudation, together with rapidly emerging information (not detailed herein) on receptors, target cells and cellular responses to the plasma-derived molecules, must be considered in any realistic model that investigates "immuno-inflammatory" mechanisms of the airway mucosa.     

Extension of a typology of alcohol dependence based on relative genetic and environmental loading

Mild, severe, and dyssocial subtypes of alcohol dependence, previously identified among Caucasian men from the Epidemiologic Catchment Area study, were also identified among Caucasian men and women with DSM-IV alcohol dependence from the National Longitudinal Alcohol Epidemiologic Survey (n = 2,703; 1,746 respectively). These subtypes were not identified among African American and Hispanic American men or women with DSM-IV alcohol dependence. Among Caucasians with alcohol dependence, the subtypes were characterized by differential loading on three dimensions: genetic, general environmental, and dyssocial environmental symptom scales developed in a prior twin study. The mild subtype (60% of men and 66% of women) was distinguished by low mean scores on all three scales; the dyssocial subtype (24% of men and 20% of women) by low mean genetic and general environmental scores but high mean dyssocial environmental scores; and the severe subtype (16% of men and 14% of women) by high scores on the genetic and general environmental scales. These subtypes also showed the expected distinctions in clinical characteristics. The severe subtype showed greater comorbid drug dependence and major depression, more treatment seeking, and a higher prevalence of parental alcoholism. The severe subtype also showed significantly greater genetic influence adjusted for overall severity of alcohol dependence (genetic ratio). Only the severe subtype showed a pattern of scale scores and clinical characteristics suggestive of substantial genetic influence. The present study indicates a robustness of the typology originally developed among DSM-III alcohol-dependent Caucasian men by empirical extension of the subtypes to a different sample of Caucasian men and, separately, Caucasian women. The use of this typology may aid in distinguishing between Caucasian alcohol-dependent individuals on the basis of relative genetic influence, enabling genetic, behavioral, and epidemiological investigations to reduce genetic or environmental "noise" and better focus on specific aspects of alcohol dependence.     

Effect of fenfluramine on food intake, mood, and performance of humans living in a residential laboratory

Five male and four female normal weight research volunteers, participating in 13-day residential studies, received oral fenfluramine (20, 40 mg) or placebo at 09:30 and 17:00. Food intake, performance, and subjective ratings were measured throughout the day. Carbohydrate intake was manipulated by providing lunch meals high (males: 120 g; females: 80 g) or low (males: 25 g; females: 16 g) in carbohydrate on 8 days; on the remaining days subjects self-selected lunch. Total caloric intake (approximately 2800 Kcal) did not differ among the low- and high-carbohydrate, and self-selected lunch conditions when subjects received placebo, indicating caloric compensation. Total carbohydrate intake was significantly less, however, when subjects consumed the low-carbohydrate lunch compared to the other lunch conditions. Fenfluramine significantly decreased total caloric intake (approximately 500 kcal) by decreasing meal size, not number, only when subjects consumed the low-carbohydrate lunch. Fenfluramine was only an effective anorectic drug when subjects consumed a lunch with fewer calories and a lower carbohydrate:protein ratio than self-selected baseline. Also, fenfluramine improved performance on a range of computer tasks and increased ratings of "Alert," "Friendly," and "Talkative," while decreasing ratings of "Tired" and "Irritable."