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51.
The integration of Fas/Apo-1 (CD95) by Fas ligand or anti-Fas antibody induces apoptosis, and this system plays a pivotal role for the lysis of target cells by cytotoxic T lymphocytes. Fas-mediated apoptosis is also increased by a prior incubation of Fas-bearing cells with interferon(IFN)-. Interleukin-1- converting enzyme (ICE) and/or CPP32, or other members of ICE family act as direct cell death executors downstream of this mechanism, and a tetrapeptide inhibitor of these cysteine proteases blocks Fas-mediated apoptosis. In this study, we examined the effect of IFN- on Fas-mediated apoptosis in ACHN cells. IFN- augmented apoptosis in a dose dependent manner and reached a plateau at 400 U/ml when exposed for 48 h before the end of culture. The kinetics revealed a significant increase in apoptosis after 24 h. Exposing ACHN cells to IFN- increased pro-ICE expression accompanied with a decrease of pro-CPP32. These results suggest that direct enhancement of ICE expression and/or upregulation of conversion of pro-CPP32 to active form increases Fas-mediated apoptosis by IFN- in ACHN cells.  相似文献   
52.
A set of 300 vaginal smears was interpreted by 13 cytologists from six European laboratories, who were requested to report inadequate and suboptimal smears. the set had been appropriately seeded to reach approximately 10% inadequate and 20% suboptimal smear frequency. According to the majority report, 230 smears were classified as adequate (76.7%), 43 as inadequate (14.3%), and 27 as suboptimal (9.0%). Agreement with the majority report ranged from 52% to 91% (average 78%). Kappa statistics for reporting inadequate smears showed a high level of agreement for five cytologists, and fair to good agreement for eight. In contrast, K statistics for reporting suboptimal smears showed fair to good agreement with the majority report only in five instances, whereas agreement was poor for eight cytologists. ‘Inadequate smear’rates may be used to compare the quality of smears received in different laboratories, as there is a high level of agreement among cytologists as to what constitutes an inadequate smear. However, this is not true for ‘suboptimal smear’ rates, and more precise reporting criteria must be defined and tested if an intermediate category is to be retained to report poor quality smears: more precise reporting criteria must be defined and tested if an intermediate category is to be retained to report poor quality smears. Une série de 300 frottis cervico-vaginaux a été examinee par 13 cytologistes appartenant à 6 laboratoires européens auxquels il avait été demandé de signaler les frottis inadéquats et sub-optimaux. Cette série de frottis a été constituée de façon à atteindre approximativement une fréquence de 10% de frottis inadéquats et de 20% de frottis sub-optimaux. Si I'on considère les réponses faites par la majorité des cytologistes, 230 frottis ont été classés adéquats (76,7%), 43 classés inadéquats (14,3%), et 27 comme sub-optimaux (9,0%). La concordance avec les réponses de la majorité des cytologistes varie de 52%à 91% (moyenne 78%). Concernant le signalement des frottis inadéquats, le test Kappa est excellent pour 5 cytologistes et moyen à bon pour les 8 autres. Au contraire, pour le signalement des frottis sub-optimaux, le test Kappa montre un accord moyen à bon avec les réponses de la majorité seulement dans 5 cas alors que I'agrément est mauvais pour 8 cytologistes. La fréquence des frottis inadéquats peut être utilisée pour comparer la qualité de prélèvement du frottis dans différents laboratoires, alors que les comparaisons de la fréquence des frottis sub-optimaux sont discutables, du fait d'un biais de diagnostic évident. Des critéres plus précis pour le signalement des frottis sub-optimaux doivent étre définis et testés si cette catégorie doit être retenue pour rapporter les frottis de mauvaise qualité. 300 Abstriche wurden durch 13 Zytologen aus 6 europäischen Laboratorien hinsichtlich der Kriterien ‘unbrauchbar’und ‘suboptimal’beurteilt. Die Sammlung enthielt etwa 10% unbrauchbare und 20% suboptimale Präparate. Mehrheitlich wurden 230 Abstriche als auswertbar (76,7%), 43 als unbrauchbar (14,3%) und 27 als suboptimal (9,0%) bezeichnet. Die Statistik ergab hinsichtlich der unbrauchbaren Abstriche eine ausgezeichnete Ûbereinstimmung für 5 und eine gute Ûbereinstim-mung für 8 Zytologen. Demgegenüber zeigte sich hinsichtlich der Beurteilung suboptimal nur 5mal eine gute und 8mal eine schlechte Ûbereinstimmung. Damit ist die Kategorie suboptimal ungeeignet zum Vergleich und bedarf einer genaueren Definition wenn sie für die Qualitätskontrolle eingesetzt werden soll.  相似文献   
53.
A retrospective study of oesophageal cytopathology at the Hospital de Clínicas de Porto Alegre (HCPA), RS, Brazil, from 1989 to 1992 was made to assess the sensitivity, specificity, predictive values and accuracy of endoscopic cytology and biopsy; and study the correlation between cytopathological and histopathological diagnosis. Specimens from 94 patients were available for review. The final diagnosis was based on surgical pathology and follow up. The 81 patients with cancer of the oesophagus had the following sex distribution: 64 males and 17 females (a 3.7–1 ratio). No tumour was found in 13 patients. The following conclusions were made: (i) there is excellent correlation between cytology and histology in oesophageal lesions sampled by endoscopy; (ii) a correct positive cytologic report was obtained in 77 (95%) of the 81 proven oesophageal cancers; a false-negative or unsatisfactory result was given in four patients. A false-positive diagnosis of cancer was not made. There were 13 true-negative reports. These findings result in a sensitivity of 95% with 95% confidence intervals (CI) of 90.26–99.74%; a specificity of 100% (CI of 98.5–100%); a positive predictive value of 100% (CI of 99.3–100%); a negative predictive value of 76% (CI of 55.7–96.3%); (iii) a correct positive histological report was obtained in 67 (83%) of the 81 proven oesophageal cancers; a false-negative or unsatisfactory result was given in 14 patients. A false-positive diagnosis of cancer was not made. There were 13 true-negative reports. These findings result in a sensitivity of 83% with 95% CI of 74.82–91.18%; a specificity of 100% (CI of 98.5–100%); a positive predictive value of 100% (CI of 99.25–100%); a negative predictive value of 48% (CI of 29.16–64.84%); (iv) of 81 patients with proven cancer, in 79 (98%) at least one of the methods was positive. In only two patients with cancer were both methods negative. These findings result in a combined sensitivity of 98% (CI of 94.92–100%); a specificity of 100% (CI of 98.5–100%); a positive predictive value of 100% (CI of 99.31–100%); and a negative predictive value of 87% (CI of 70–100%). Our series confirms the value of the combined use of cytology and biopsy for the investigation of oesophageal lesions. However, it should be remembered that even with the combined use of cytology and biopsy there are some tumours that will be negative by both procedures: we had only two such cases, confirming the rarity of such an event.  相似文献   
54.
Summary Laminin synthesis and deposition are concomitant with the development of a basal lamina between the human epidermis and the underlying dermis. One of the challenges in tissue engineering of human epidermal models is to develop substrates and conditions that encourage the development of a basement membrane. The purpose of this study was to determine if actin filaments and/or microtubules are involved in the synthesis/secretion of laminin by normal human epidermal keratinocytes (NHEK)in vitro. NHEK synthesize and secrete laminin subunits B1, B2, and M but little, if any, of laminin subunit A. Data indicate that disruption of microfilaments by the destabilizing agent, cytochalasin D, had no apparent effect on the relative synthesis rates of most cytosolic proteins as, revealed by one-dimensional sodium dodecyl sulfate (SDS) gel electrophoresis. This drug, however, increased laminin B2 synthesis several fold over untreated controls. This enhanced synthetic rate was independent of the type of collagen, matrix on which the NHEK were grown. Similar increases in synthesis of the M and B1 laminin chains were not observed. To determine if this increase in synthesis lead to increases in laminin B2 secretion, laminin B2 was immunoprecipitated from both the apical and basal domains of NHEK cells grown on microporous membranes. While more laminin B1, B2, and M were secreted basally than apically, an observation consistent with laminin’s role in basal lamina formation, cytochalasin D had no apparent effect on either basal or apical laminin B2 secretion. Experiments with the microtubule destabilizer, nocodazole, showed no similar effects on laminin synthesis and/or secretion. We conclude that (a) disruption of the actin network in NHEK selectively increases the synthesis of laminin B2, (b) the secretion of laminin B2 from NHEK cells is not governed by either the microfilamentous cytoskeleton or the amount of laminin synthesized by NHEK, and (c) disruption of the microtubular network does not alter laminin synthesis or secretion.  相似文献   
55.
The Nucleotide Sequence of Human Acylamino Acid-Releasing Enzyme   总被引:3,自引:0,他引:3  
The nucleotide sequence of a cDNA coding for the human acylaminoacid-releasing enzyme (AARE, also known as acylpeptide hydrolase)[EC 3.4.19.1] subunit has been determined. The amino acid sequenceof human AARE subunit deduced from its cDNA nucleotide sequenceshowed a high degree of identity (91.5%) with both the correspondingproteins from the pig and the rat. The AARE cDNA shows 99.2%identity with a 3.3 kb cDNA transcribed from a locus (DNF15S2)on the short arm of human chromosome 3, whose deletion is associatedwith small cell lung cancer, taking into consideration thatthe sequence of the 3.3-kb cDNA previously reported was causedby misreading.  相似文献   
56.
Muscular activity converts chemical energy into useful work and metabolism restores muscle to its original state. This essay explores the organization and interactions of the regulatory system(s) which allow this energy balance to occur. The term energy balance is used in a biochemical rather than a thermodynamic sense—concerned not with deductions from the physical principles of thermodynamics, but rather with those enzymatic processes which nature evolved and which operate at remarkably fixed stoichiometry. Energy balance is a statement of conservation of energy put into biochemical observables.31P NMR spectroscopy is one of the most useful techniques for investigating these questions quantitatively under physiological conditionsin vivo. The author (1) describes the rules or principles of biochemical energy balance; (2) discusses sample results from human muscle to demonstrate its use in studying this class of questions; (3) presents a simple model of integrated cellular respiration to demonstrate its sufficiency to account for the main observations.  相似文献   
57.
The dose response effect of a new adenosine analogue, GR 79236 (N-[1S trans-2-hydroxycyclopentyl] adenosine) upon insulin sensitivity was examined in human adipocytes. The influence of adenosine upon insulin sensitivity for suppression of lipolysis and stimulation of glucose transport was examined. Removal of adenosine by use of adenosine deaminase stimulated lipolysis to the same extent as did 10–9 M noradrenaline. GR79236 brought about dose dependent inhibition of lipolysis with half-maximal effect at 11.3±7.8×10–9 M. When lipolysis was stimulated by noradrenaline alone the subsequent inhibition of lipolysis brought about by GR79236 was significantly greater than that of insulin. To examine adenosine effects on the insulin signalling pathway separately from those on lipolysis, the insulin sensitivity of glucose transport was examined. Removal of adenosine brought about a small but significant increase in the concentration of insulin required for half-maximal stimulation of glucose transport. Adenosine agonists offer promise as new agents for the modulation of metabolism in diabetes and other states of insulin resistance.  相似文献   
58.
Summary Primary and passaged cultures of normal colon epithelial cells, derived from human fetuses (13 to 17 wk of conceptual age) have been established. These cultures have been passaged 16 times thus far. The cultures have been initiated and maintained in medium consisting of 50% Dulbecco's minimum essential medium and 50% Ham's F12 medium and supplemented with antibiotics (penicillin, 100 U/ml; streptomycin, 100 μg/ml); ascorbic acid, 40 μg/ml;l-isoleucine, 50 μg/ml; epidermal growth factor, 20 ng/ml; insulin, 5 μg/ml; cholera toxin, 5 ng/ml; transferrin, 1 μg/ml; fetal bovine serum (10%); and HEPES, 25 mM final concentration, and incubated at 37°C in humidified gas containing 5% CO2: 95% air. The cellular and subcellular characteristics of primary and passaged cultures were defined using light microscopy and scanning and transmission electron microscopy. The cells exhibited microvilli on cell surfaces and showed junctional complexes and interdigitations between cells. Indented nuclei with dense chromatin and marginated heterochromatin, numerous mitochondria, rough endoplasmic reticulum, polysomes, and extensive Golgi zones were conspicuous. Also, periodic acid Schiff's reagent-positive staining of the cells suggests the active synthesis of complex mucopolysaccharides in the cytoplasm. This study was supported by USPHS Grant CA-30185 from the National Large Bowel Cancer Project, National Cancer Institute.  相似文献   
59.
The rotational freedom of tryptophan residues in protein-ligand complexes was studied by measuring steady-state fluorescence anisotropies under conditions of oxygen quenching. There was a decrease in the oxygen bimolecular quenching constant upon complexation of trypsin and alpha-chymotrypsin with proteinaceous trypsin inhibitors, of lysozyme with N-acetylglucosamine (NAG) and di(N-acetyl-D-glucosamine) ((NAG)2) and of hexokinase with glucose. Binding of the bisubstrate analogue N-phosphonacetyl-L-aspartate (PALA) to aspartate transcarbamylase (ATCase) and binding of biotin to avidin resulted in increased oxygen quenching constants. The tryptophan of human serum albumin (HSA) in the F state was more accessible to oxygen quenching than that in the N state. With the exception of ATCase, the presence of subnanosecond motions of the tryptophan residues in all the proteins is suggested by the short apparent correlation times for fluorescence depolarization and by the low apparent anisotropies obtained by extrapolation to a lifetime of zero. Complex formation evidently resulted in more rigid structures in the case of trypsin, alpha-chymotrypsin and lysozyme. The effects of glucose binding on hexokinase were not significant. Binding of biotin to avidin resulted in a shorter correlation time for the tryptophan residues. The N --> F transition in HSA resulted in a more rigid environment for the tryptophan residue. Overall, these changes in the dynamics of the protein matrix and motional freedom of tryptophan residues due to complex formation and subsequent conformational changes are in the same direction as those observed by other techniques, especially hydrogen exchange. Significantly, the effects of complex formation on protein dynamics are variable. Among the limited number of cases we examined, the effects of complex formation were to increase, decrease or leave unchanged the apparent dynamics of the protein matrix.  相似文献   
60.
Photoaffinity labeling techniques have recently demonstrated that mammalian β1- and β2-adrenergic receptors reside on peptides of Mr 62 000–64 000. These receptor peptides are susceptible to endogenous metalloproteinases which produce peptides of Mr 30 000–55 000. Several proteinase inhibitors markedly attenuate this process, specifically EDTA and EGTA. In this study we investigated the functional significance of this proteolysis (and its inhibition) in the β2-adrenergic receptor-adenylate cyclase system derived from rat lung membranes. Membrane preparations containing proteolytically derived fragments of the receptor of Mr 40000–55 000 are fully functional with respect to their ability to bind β-adrenergic antagonist radioligands such as [3H]dihydroalprenolol and β-adrenergic antagonist photoaffinity reagents such as p-azido-m-[125I]iodobenzylcarazolol. They retain the ability to form a high-affinity, agonist-promoted, guanine nucleotide-sensitive complex thought to represent a ternary complex of agonist, receptor and guanine nucleotide regulatory protein. Nonetheless, after proteolysis, GTP is less able to revert this high-affinity receptor complex to one of lower affinity, and all aspects of adenylate cyclase stimulation are reduced. In addition, the functional integrity of the N protein in membranes prepared without proteinase inhibitors is reduced as assessed by reconstitution studies with the cyc[su− variant of S49 lymphoma cell membranes. These results suggest that endogenous proteolysis does not directly impair the ability of β-adrenergic receptors to either bind ligands or interact with the guanine nucleotide regulatory protein. However, they imply that endogenous proteolysis likely impairs the functionality of other components of the adenylate cyclase system, such as the nucleotide regulatory protein.  相似文献   
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