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A Haas

Publications and source records attributed to A Haas.

At least 37 records · Page 2Linked to original sources
PubMed2001

Use of recombinant mitogillin for improved serodiagnosis of Aspergillus fumigatus-associated diseases.

During human infection, Aspergillus fumigatus secretes a 18-kDa protein that can be detected as an immunodominant antigen in the urine of infected patients. Recently, this protein was shown to be mitogillin, a ribotoxin that cleaves a single phosphodiester bond of the 29S rRNA of eukaryotic ribosomes. We proved the immunogenic capacity of mitogillin in a rabbit animal model, indicating its usefulness as an antigen for serological diagnosis of invasive aspergillosis. The mitogillin gene from A. fumigatus was transferred from plasmid pMIT+ to expression vector pQE30 and expressed in Escherichia coli as a fusion protein. Purified recombinant mitogillin was recognized by serum immunoglobulin G (IgG) of polyclonal rabbit sera that were obtained by immunization with purified native mitogillin. Consequently, we developed an enzyme-linked immunosorbent assay for detection of IgG, IgM, and IgA antibodies to recombinant mitogillin. In serum samples of patients suffering from aspergilloma (AO; n = 32), invasive pulmonary aspergillosis (IPA; n = 42), or invasive disseminated aspergillosis (IDA; n = 40), a good correlation of production of IgG antibody against mitogillin and clinical disease was observed (for patients with AO, 100% [32 of 32] were positive; for patients with IPA, 64% [31 of 42] were positive; for patients with IDA, 60% [24 of 40] were positive). In contrast, positive titers for serum IgG and IgM antibodies against mitogillin were found in only 1.3% of the serum samples of healthy volunteers and positive titers for IgA antibody were found in only 1.0% of the serum samples of healthy volunteers (n = 307; specificity = 95.4%). These results indicate that recombinant mitogillin expressed in E. coli can be used for improvement of the serodiagnosis of A. fumigatus-associated diseases.

Adolescent↗
PubMed2001

Hyperhomocyst(e)inaemia, but not MTHFR C677T mutation, as a risk factor for non-arteritic ischaemic optic neuropathy.

BACKGROUND/AIMS: Hyperhomocyst(e)inaemia has been identified as a strong risk factor for stroke, myocardial infarction, and deep vein thrombosis. A point mutation of methylene tetrahydrofolate reductase (MTHFR C677T) has been associated with increased plasma homocyst(e)ine levels. To investigate whether hyperhomocyst(e)inaemia and/or MTHFR C677T mutation are associated with non-arteritic ischaemic optic neuropathy (NAION), a case-control study including 59 consecutive patients with NAION and 59 controls matched for age and sex was performed. METHODS: Fasting plasma homocyst(e)ine levels, MTHFR C677T genotypes, and plasma levels of folate and vitamin B-12 were determined. RESULTS: Mean plasma homocyst(e)ine levels were significantly higher in patients than in controls (11.8 (SD 5.7) micromol/l v 9.8 (2.5) micromol/l, p = 0.02). The odds ratio for patients with homocyst(e)ine levels exceeding the 95th percentile of control homocyst(e)ine levels was 5.8 (95% CI 1.5-21.4). Mean plasma folate levels were significantly lower in patients than in controls (4.3 (1.7) ng/ml v 5.5 (1.9) ng/ml, p = 0.001), whereas plasma vitamin B-12 levels did not differ significantly. Prevalence of the MTHFR C677T mutation was not significantly increased in patients with NAION compared with controls. CONCLUSION: These results suggest that hyperhomocyst(e)inaemia, but not MTHFR C677T mutation is associated with NAION. Determination of plasma homocyst(e)ine levels might be of diagnostic value in patients with NAION.

Aged↗
PubMed2001

Three-dimensional reconstruction of the anal striated musculature in a human fetus.

Conflicting opinions in the recent literature indicate that the morphological organization and function of the anorectal continence organ has, up to now, not been clearly understood. But a clear imagination of the spatial arrangements of this compound muscle system is of clinical relevance for the pediatric surgeon performing reconstructive surgery. We analyzed 18-microm sections of the pelvic region of 4 human fetuses in order to describe the individual components of this muscle complex. A series of 630 Azan-stained sections was the base for the computer-assisted 3D reconstruction of the levator ani and the external sphincter complex in a male human fetus (14th week p.c.). In this context, special attention was paid to the intermediate muscle layer of the puborectalis which develops ventrally from the funnel-shaped levator ani and joins the tripartite ring system of the sphincter muscle dorsally. Our findings lead to a clear imagination of the spatial arrangement of this intermediate layer and characterize the anorectal muscle complex as an integrated ensemble in which the puborectalis holds a key position.

Anal Canal↗
PubMed2001

Development of an experimental paradigm in which to examine human learning using two computer-based formats.

Computer-based self-instructional programs are frequently promoted as means to augment or replace the traditional anatomy curricula taught in medical schools. These programs may range from static slide shows to fully immersive virtual environments. However, the impact of these learning technologies on knowledge acquisition, and their comparative cost/benefit to education remain unclear. As a consequence, we are embarking on a series of experiments to compare knowledge acquisition and the meaningful use of information among students who are learning anatomy using one of two different computer-based self-instructional formats. These studies will be based on a specially developed learning module on basic lung anatomy; they will utilize a variety of assessment tools to measure factual knowledge, conceptual understanding of spatial-anatomic relationships, and the ability to apply newly acquired knowledge of anatomy to clinical problem-solving scenarios. The primary object of this paper is to describe the design and development of the underlying test module and to outline the two computer-based formats that will be evaluated. The virtual reality (VR) environment, UCSD's Anatomic VisualizeR, provides dynamic access to 3-dimensional polygonal models of the lesson content and supports student-centered exploration and learning. The multimedia environment, Microsoft PowerPoint, provides a structured presentation of the lesson content and illustrates important anatomic structures through the use of 2-dimensional images derived by screen captures of models available in the VR learning module. This paper also provides an overview of the first experiment in the series, a pilot study using first-year medical students without previous participation in a medical school anatomy curriculum. For this study, students will be prospectively randomized into two groups, each group learning the lung anatomy lesson using one of the computer-based formats described. Immediate knowledge retention will be measured by asking students to complete the assessment instrument immediately after completing their learning module. The results of the pilot study will be used to refine and improve the design of the remainder of studies planned in this experimental series.

Anatomy↗
PubMed2000

Variation in anuran embryogenesis: differences in sequence and timing of early developmental events.

Comparative embryology of closely related species can shed light on the evolution of developmental processes. An important mechanism in the evolution of developmental processes, which can lead to significant changes in larval or adult form, is variation in the sequence and timing of developmental events. We compared the development of 12 species of anurans, including a wide taxonomic range as well as a number of congeneric species. The comparison consisted of monitoring a series of external morphological markers and histological markers. For each species we noted the timing of each of the markers, using a uniform parameter of normalized time. We compared the normalized time of each of these events among the species, as well as the sequence of the events. Our analysis revealed many differences in sequence and in timing of developmental events. We mapped these differences on a cladogram of the studied species, using sequence units as discrete characters. The differences do not seem to be connected to the phylogenetic relations between the species or to any obvious ecological factors. We suggest a hypothetical ancestral sequence of developmental events, and discuss the possible factors that could have caused the observed variations from the ancestral sequence.

Animals↗
PubMed2000

Sequential action of two GTPases to promote vacuole docking and fusion.

Homotypic vacuole fusion occurs by sequential priming, docking and fusion reactions. Priming frees the HOPS complex (Vps 11, 16, 18, 33, 39 and 41) to activate Ypt7p for docking. Here we explore the roles of the GDP and GTP states of Ypt7p using Gdi1p (which extracts Ypt7:GDP), Gyp7p (a GTPase-activating protein for Ypt7p:GTP), GTPgammaS or GppNHp (non-hydrolyzable nucleotides), and mutant forms of Ypt7p that favor either GTP or GDP states. GDP-bound Ypt7p on isolated vacuoles can be extracted by Gdi1p, although only the GTP-bound state allows docking. Ypt7p is converted to the GTP-bound state after priming and stably associates with HOPS. Gyp7p can cause Ypt7p to hydrolyze bound GTP to GDP, driving HOPS release and accelerating Gdi1p-mediated release of Ypt7p. Ypt7p extraction does not inhibit the Ca(2+)-triggered cascade that leads to fusion. However, in the absence of Ypt7p, fusion is still sensitive to GTPgammaS and GppNHp, indicating that there is a second specific GTPase that regulates the calcium flux and hence fusion. Thus, two GTPases sequentially govern vacuole docking and fusion.

Calcium↗
PubMed2000

Essential role of Glu-C66 for menaquinol oxidation indicates transmembrane electrochemical potential generation by Wolinella succinogenes fumarate reductase.

Quinol:fumarate reductase (QFR) is a membrane protein complex that couples the reduction of fumarate to succinate to the oxidation of quinol to quinone, in a reaction opposite to that catalyzed by the related enzyme succinate:quinone reductase (succinate dehydrogenase). In the previously determined structure of QFR from Wolinella succinogenes, the site of fumarate reduction in the flavoprotein subunit A of the enzyme was identified, but the site of menaquinol oxidation was not. In the crystal structure, the acidic residue Glu-66 of the membrane spanning, diheme-containing subunit C lines a cavity that could be occupied by the substrate menaquinol. Here we describe that, after replacement of Glu-C66 with Gln by site-directed mutagenesis, the resulting mutant is unable to grow on fumarate and the purified enzyme lacks quinol oxidation activity. X-ray crystal structure analysis of the Glu-C66-->Gln variant enzyme at 3.1-A resolution rules out any major structural changes compared with the wild-type enzyme. The oxidation-reduction potentials of the heme groups are not significantly affected. We conclude that Glu-C66 is an essential constituent of the menaquinol oxidation site. Because Glu-C66 is oriented toward a cavity leading to the periplasm, the release of two protons on menaquinol oxidation is expected to occur to the periplasm, whereas the uptake of two protons on fumarate reduction occurs from the cytoplasm. Thus our results indicate that the reaction catalyzed by W. succinogenes QFR generates a transmembrane electrochemical potential.

Amino Acid Sequence↗
PubMed2000

Morphologic characteristics of disciform scarring after radiation treatment for age-related macular degeneration.

OBJECTIVE: To investigate the influence of radiation therapy on the development of disciform lesions in patients with age-related macular degeneration (AMD). DESIGN: A prospective, nonrandomized, comparative trial (patient self-controlled). PARTICIPANTS: Forty eyes with exudative AMD involving the central fovea in 40 consecutive patients were enrolled in this study. INTERVENTION: Radiation was administered to the posterior pole with an 8-mV photon beam from a linear accelerator. A dose of 14.4 Gy, 1.8 Gy per day, five fractions per week was delivered through a single port. MAIN OUTCOME MEASURES: The visual acuity and the morphologic characteristics, demonstrated by fundus photography, fluorescein, and indocyanine green angiography, were investigated before treatment and every 3 months after treatment over a period of 24 months. In 10 patients with bilateral disease the disciform lesions were compared. RESULTS: Twenty five patients could be followed regularly over the period of 24 months. The disciform lesions occurring after radiation were classified in three types. Type I (10 patients) was characterized by being smaller than 2 DD in size, with little fibrotic tissue underneath the retina, but pronounced retinal pigment epithelial changes. Type II (seven patients) showed extensive growth of the choroidal neovascularization (CNV) extending to and beyond the arcades with angiographically active loops in the peripheral parts. Eight patients had type III lesions develop characterized by a size greater than 2 DD but fewer than 6 DD and by a different amount of fibrotic tissue, hemorrhage, and lipid. Type I scarring was significantly associated with occult CNV without pigment epithelial detachments, whereas type II scarring was associated with classic CNV at the initial presentation (P<0.05). CONCLUSIONS: Although no severe side effects have been reported after radiation therapy for AMD, a subgroup of patients may experience extensive growth of CNV after radiation, causing greater functional damage than occurs spontaneously.

Aged↗
PubMed2000

A method to purify bacteria-containing phagosomes from infected macrophages.

When small particles, such as microorganisms, are taken up by macrophages, they are wrapped with a portion of the host cell plasma membrane and ingested, creating a new organelle, the phagosome. This phagosome matures stepwise as newly formed endosomes do, finally forming a phagolysosome, a process that contributes to killing of ingested microbes and to the presentation of microbial antigens on the surface of the phagocyte. Some pathogenic bacteria, however, reprogram the phagocytic cell in such a way that the phagosome will either be arrested in an early stage of maturation or will be diverted and create an unusual, novel phagosomal compartment. To study the molecular processes that underly biogenesis of bacteria-containing phagosomes, we have established a method to isolate and to biochemically analyse bacteria- containing phagosomes. This method consists of mechanical lysis of infected macrophages, production of a postnuclear supernatant followed by fractionation in a discontinuous sucrose density gradient, separation through a Ficoll cushion, and by a final concentration step. These phagosome preparations contain very little endosomal or lysosomal contamination (the organelles of most concern when studying phagosome biogenesis) and very little Golgi- and plasma membrane-derived contamination, but do contain some mitochondrial and ER contamination. This method could also be used to study bacterial factors (proteins, RNA) produced while in phagosomes.

Afipia↗
PubMed2000

Optical coherence tomography (OCT) in acute macular neuroretinopathy.

PURPOSE: To evaluate the value of ocular coherence tomography (OCT) concerning diagnosis and pathogenesis of acute macular neuroretinopathy. METHODS: A 33-year old woman complained of sudden onset of central scotomas in her right eye because of acute macular neuroretinopathy. We performed a direct ophthalmoscopy, a visual field testing, a fluorescein angiography (FA) a multifocal ERG (mf-ERG) and an OCT. RESULTS: We found typical paracentral scotoma in visual field testing, a normal FA and mf-ERG in her right eye. In OCT there was a band of higher reflectivity (115 microm) overlying an intact band corresponding to the retinal pigment epithelium (RPE)/ choriocapillaris complex. Retinal thickness was within the normal range. CONCLUSION: OCT can be an additional valuable tool in acute macular neuroretinopathy as it is a disease with discrete pathology and often normal results in other diagnostic tests.

Acute Disease↗
PubMed2000

Phosphatidylinositol 4,5-bisphosphate regulates two steps of homotypic vacuole fusion.

Yeast vacuoles undergo cycles of fragmentation and fusion as part of their transmission to the daughter cell and in response to changes of nutrients and the environment. Vacuole fusion can be reconstituted in a cell free system. We now show that the vacuoles synthesize phosphoinositides during in vitro fusion. Of these phosphoinositides, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)) are important for fusion. Monoclonal antibodies to PI(4,5)P(2), neomycin (a phosphoinositide ligand), and phosphatidylinositol-specific phospholipase C interfere with the reaction. Readdition of PI(4, 5)P(2) restores fusion in each case. Phosphatidylinositol 3-phosphate and PI(3,5)P(2) synthesis are not required. PI(4,5)P(2) is necessary for priming, i.e., for the Sec18p (NSF)-driven release of Sec17p (alpha-SNAP), which activates the vacuoles for subsequent tethering and docking. Therefore, it represents the kinetically earliest requirement identified for vacuole fusion so far. Furthermore, PI(4,5)P(2) is required at a step that can only occur after docking but before the BAPTA sensitive step in the latest stage of the reaction. We hence propose that PI(4,5)P(2) controls two steps of vacuole fusion.

Membrane Fusion↗
PubMed2000

Phagosome acidification has opposite effects on intracellular survival of Bordetella pertussis and B. bronchiseptica.

Bordetella pertussis is readily killed after uptake by professional phagocytes, whereas its close relative Bordetella bronchiseptica is not and can persist intracellularly for days. Phagocytosis of members of either species by a mouse macrophage cell line results in transport of the bacteria to a phagosomal compartment positive for the lysosome-associated membrane protein 1, the protease cathepsin D, and the late endosomal vacuolar proton-pumping ATPase but negative for the early endosome antigen 1 and the early endosomal transferrin receptor. In addition, we demonstrate that Bordetella-containing phagosomes rapidly acidify to pH 4.5 to 5.0. Taken together, these data demonstrate that Bordetella-containing phagosomes rapidly mature to an acidic late endosomal/lysosomal compartment. Following up on this observation, we determined that B. pertussis does not survive in bacterial growth media adjusted to a pH of 4.5, whereas this pH has only minor effects on the growth of B. bronchiseptica. Raising the intracellular pH in infected macrophages by the addition of bafilomycin A(1), ammonium chloride, or monensin increases the survival of acid-sensitive B. pertussis but, surprisingly, decreases that of acid-tolerant B. bronchiseptica. In summary, we hypothesize that the differential survival of B. pertussis and B. bronchiseptica in macrophages is, at least in part, due to the differences in their acid tolerance.

Animals↗