Determination of the enzymes responsible for activation of the heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline in the human breast

The aim of this work was to study the effects of benzalkonium chloride (BAC) treatment on the small intestine and its functioning in rats surgically prepared with Thiry-Vella intestinal loop. The loops were treated with either BAC, which ablated much of the myenteric plexus and extrinsic innervation, or with physiological saline (SAL). In vivo drinking experiments were performed to examine the effect on fluid intake and behavioral indices of distending the loop with a balloon. Spontaneous motility and its changes induced by acetylcholine (ACh) and histamine (His) were studied on isolated stripes in vitro. Finally, samples from the loops were examined histologically. Though reduction of the cell number was less than expected and no differences of the thickness of the muscular layer between the two groups was observed, BAC treatment altered the pattern of spontaneous activity and also the sensitivity to ACh and His in isolated stripes. In vivo distension of the SAL-treated loops reduced fluid intake and produced signs of aversivity; these effects were absent in the BAC-treated group. Our results show that despite the differences in the degree of ablation from those obtained by others, BAC treatment can be used to study the mechanisms underlying the effects of the enteral stimuli on the behavior.     

Relevance of the D13 region to the function of the skeletal muscle chloride channel, ClC-1

Although hydropathy analysis of the skeletal muscle chloride channel protein, ClC-1, initially predicted 13 potential membrane spanning domains (D1 to D13), later topological studies have suggested that domain D4 is extracellular and that D13, conserved in all eukaryotic ClC channels, is located within the extensive cytoplasmic tail that makes up the carboxyl terminus of the protein. We have examined the effect of deleting D13 (DeltaD13) and the function of the carboxyl tail by removing the final 72 (fs923X), 100 (fs895X), 125 (L869X), 398 (N596X), and 420 (Q574X) amino acids from rat ClC-1. Appropriate cDNA constructs were prepared and expressed using the baculovirus Sf9 insect cell system. Patch clamp analysis of chloride currents in Sf9 cells showed that only relatively insubstantial changes could be attributed to the expressed fs923X, fs895X, and DeltaD13 mutants compared with wild type rat ClC-1. For N596X and Q574X, however, adequate mRNA could be detected, but neither patch clamp nor polyacrylamide gel electrophoresis showed corresponding protein production. By contrast, expression of L869X was demonstrable by polyacrylamide gel electrophoresis, but no chloride conductance attributable to it could be detected. Overall, our results indicate that the domain D13 is dispensable, as are the final 100 amino acids, but not the final 125 amino acids or more, of the carboxyl tail. Some essential region of unknown significance, therefore, appears to reside in the 18 amino acids after D13, from Lys877 to Arg894.     

Forecasting generalized epileptic seizures from the EEG signal by wavelet analysis and dynamic unsupervised fuzzy clustering

Dynamic state recognition and event-prediction are fundamental tasks in biomedical signal processing. We present a new, electroencephalogram (EEG)-based, brain-state identification method which could form the basis for forecasting a generalized epileptic seizure. The method relies on the existence in the EEG of a preseizure state, with extractable unique features, a priori undefined. We exposed 25 rats to hyperbaric oxygen until the appearance of a generalized EEG seizure. EEG segments from the preexposure, early exposure, and the period up to and including the seizure were processed by the fast wavelet transform. Features extracted from the wavelet coefficients were imputed to the unsupervised optimal fuzzy clustering (UOFC) algorithm. The UOFC is useful for classifying similar discontinuous temporal patterns in the semistationary EEG to a set of clusters which may represent brain-states. The unsupervised selection of the number of cluster overcomes the a priori unknown and variable number of states. The usually vague brain state transitions are naturally treated by assigning each temporal pattern to one or more fuzzy clusters. The classification succeeded in identifying several, behavior-backed, EEG states such as sleep, resting, alert and active wakefulness, as well as the seizure. In 16 instances a preseizure state, lasting between 0.7 and 4 min was defined. Considerable individual variability in the number and characteristics of the clusters may postpone the realization of an early universal epilepsy warning. University may not be crucial if using a dynamic version of the UOFC which has been taught the individual's normal vocabulary of EEG states and can be expected to detect unspecified new states.     

Prevention of postoperative thrombotic stroke after carotid endarterectomy: the role of transcranial Doppler ultrasound

PURPOSE: To determine the incidence of particulate embolization after carotid endarterectomy (CEA), the effect of dextran-40 infusion in patients with sustained postoperative embolization, and the impact of transcranial Doppler (TCD) monitoring plus adjuvant dextran therapy on the rate of postoperative carotid thrombosis. METHODS: Prospective study in 100 patients who underwent CEA with 6-hour postoperative monitoring using a TCD that was modified to allow automatic, intermittent recording from the ipsilateral middle cerebral artery waveform (10 minute sample every 30 minutes). An incremental dextran-40 infusion was commenced if 25 or more emboli were detected in any 10-minute period. RESULTS: Overall, 48% of patients had one or more emboli detected in the postoperative period, particularly in the first 2 hours. However, sustained embolization that required Dextran therapy developed in only five patients. In each case, the rate of embolization rapidly diminished. CONCLUSIONS: A small proportion of patients have sustained embolization after CEA, which in previous studies has been shown to be highly predictive of thrombotic stroke. Intervention with dextran reduced and subsequently stopped all the emboli in those in whom it was used and contributed to a 0% perioperative morbidity and mortality rate in this series.     

Evaluation of a new, integral, whole blood filter (RS2000) system for prestorage leucodepletion of SAG-M red cells

We utilized indirect immunocytochemistry to demonstrate the presence of growth factors and their receptors in human pre-embryos and Fallopian tubes. In pre-embryos, only transforming growth factor-alpha (TGF-alpha) and the intracellular domain of epidermal growth factor receptor (EGFR) were found at the 4-cell stage. In 8- to 14-cell pre-embryos, TGF-alpha, the intracellular and extracellular domains of EGFR, and insulin-like growth factor-I and its receptor were found. Antibodies against TGF-alpha stained all Fallopian tube specimens, while the extracellular domains of EGFR was only found in specimens from patients with either blood type A or AB. These results suggest a cross-reactivity between the extracellular domain of the EGFR and blood group antigens. Our novel demonstration of growth factor receptor staining in human pre-embryos shows that growth factor receptor localization is dependent on the developmental stage of human pre-embryos. We have also established a potentially important link between the Fallopian tube which secretes growth factors and the localization of growth factor receptors in pre-embryos. These findings are compatible with the hypothesis that tubal secretions are embryotrophic for the early development of the pre-embryo.     

Collagen fibers as a temporary scaffold for replacement of ACL in goats

ACL substitutes made of braided or plied purified collagen fibers and cross-linked with hexamethylenediisocyanate were implanted into a total of 14 adult goats to achieve resorption within 8 to 10 months. Two types of collagen fiber prostheses differing in degree of collagen purification were tested. The implants were harvested 2 to 11 months postimplantation, tested for mechanical strength, and evaluated by morphological methods. In the first group (n = 5), the less purified and less cross-linked collagen fiber ACL implant induced fast connective tissue ingrowth. At 6 months postimplantation, 40 to 60% of the collagen implant was resorbed. No studies on breaking strength were done in this group. In the second group, highly purified and more crosslinked ACL implants were less infiltrated by cells and were resorbed only by 10 to 20%. Still, the breaking strength was decreased to 10% of the original implant strength. In the second group, the fixation of the ACL implant in the bone tunnel with a bone wedge was insufficient (n = 6); however, additional fixation with metal screws was successful (n = 3). We conclude that cross-linked collagen fibers alone cannot be used as a safe ACL substitute as they quickly lose mechanical strength despite limited biodegradation.     

Human cytomegalovirus-infected cells have unstable assembly of major histocompatibility complex class I complexes and are resistant to lysis by cytotoxic T lymphocytes

Viruses which cause persistence in the naturally infected host are predicted to have evolved immune evasion mechanisms. Human cytomegalovirus (HCMV) causes significant morbidity and mortality in immunocompromised patients yet persists without clinical manifestations in seropositive individuals who have normal immune function. We report that HCMV infection in vitro impairs major histocompatibility complex class I (MHC-I) assembly accompanied by resistance to killing by cytotoxic CD8+ T lymphocytes. Pulse-chase metabolic labelling experiments show that MHC-I complexes continue to be assembled by both uninfected and HCMV-infected cells. However, MHC-I molecules are unstable in HCMV-infected cells and are rapidly broken down. Endoglycosidase H treatment of immunoprecipitates indicates that the breakdown of MHC-I complexes in HCMV-infected cells occurs primarily in a pre-Golgi compartment. Interference with normal MHC-I assembly and expression, if relevant in vivo, may have implications for the restriction of the diversity of the CD8+ cytotoxic T lymphocyte repertoire directed against HCMV antigens and may be an important mechanism of viral persistence.