Normal aging effects for frequency doubling technology perimetry.

PURPOSE: To determine the influence of normal aging on contrast sensitivity for frequency doubling technology (FDT) perimetry. METHODS: Contrast sensitivity measures were obtained for frequency-doubled stimuli (0.25 cycles per degree sinusoidal gratings undergoing 25 Hz counterphase flicker) at 17 target locations (4 per quadrant plus the central 5 degrees ) using a prototype of the Welch Allyn (Skaneateles, NY)/Humphrey Systems FDT perimeter (Humphrey Systems, Dublin, CA). A total of 407 normal subjects (761 eyes) between the ages of 15 and 85 years were tested. RESULTS: Between the ages of 15 and 60 years there was an approximately linear decrease in contrast sensitivity of 0.6 dB per decade. After the age of 70, there was a slightly greater sensitivity loss with age. There were no meaningful differences in sensitivity loss as a function of age for different visual field locations. A small but consistent reduction in contrast sensitivity (approximately 0.7 dB) was found at all visual field locations for the second eye tested that may be due to a central adaptation process. CONCLUSIONS: Normal aging effects for FDT perimetry are similar to those obtained for conventional automated perimetry, except that the FDT perimetry aging effects do not appear to be eccentricity dependent. These normative data provide a basis for establishing a statistical analysis procedure and probability plots for FDT perimetry.

The repeatability of automated and clinician refraction.

PURPOSE: Auto-refractors are used as a starting point for clinicians' refractions and in studies of refractive error. We investigated the repeatability of the Hoya AR-570 and clinician refraction. METHODS: Eighty-six subjects, aged 11 to 60 years, were recruited by mailing inquiries to 500 randomly selected patients who had received recent examinations at the University of California Optometric Eye Center. Contact lens wearers, patients with best corrected visual acuity worse than 20/30 in either eye, and patients with a history of diabetes were excluded. Each subject was examined by two clinicians during one visit. The first clinician obtained five auto-refractor readings for each eye (which were later averaged), performed a balanced subjective refraction (with spherical masking lenses in the phoropter), and repeated the automated refractor measurements. This protocol was then repeated by the second clinician. Clinicians were randomized with regard to testing order and masked to automated refractor results, each other's refractions, and previous spectacle prescriptions. RESULTS: To quantify repeatability, we used mixed model analyses of variance to estimate the appropriate variance components while accounting for the correlation among, for example, repeated measurements of the same eye. Astigmatic data were analyzed by converting into Fourier form: two cross-cylinders at axis 0 degrees (J0) and axis 45 degrees (J45). For mean spherical equivalent, the average difference between five averaged automated refractor readings, taken by two different optometrists, was +0.02 D (95% limits of agreement = -0.36 to +0.40 D). The average difference between the two optometrists' subjective refractions was -0.12 D (95% limits of agreement = -0.90 to +0.65 D). The 95% limits of agreement for the automated refractor were about half those of the clinician for both astigmatic terms (J0 and J45) and for all comparisons. CONCLUSIONS: Automated refraction is more repeatable than subjective refraction and therefore more appropriate for studies of myopia progression.

An efficient route to human bispecific IgG.

Production of bispecific IgG (BsIgG) by coexpressing two different antibodies is inefficient due to unwanted pairings of the component heavy and light chains. To overcome this problem, heavy chains were remodeled for heterodimerization using engineered disulfide bonds in combination with previously identified "knobs-into-holes" mutations. One of the variants, S354C:T366W/Y349'C:T366'S:L368'A:Y407++ +'V, gave near quantitative (approximately 95%) heterodimerization. Light chain mispairing was circumvented by using an identical light chain for each arm of the BsIgG. Antibodies with identical light chains that bind to different antigens were identified from an scFv phage library with a very restricted light chain repertoire for the majority (50/55) of antigen pairs tested. A BsIgG capable of simultaneously binding to the human receptors HER3 and cMpI was prepared by coexpressing the common light chain and corresponding remodeled heavy chains followed by protein A chromatography. The engineered heavy chains retain their ability to support antibody-dependent cell-mediated cytotoxicity as demonstrated with an anti-HER2 antibody.

Role of arginine-43 and arginine-69 of the Hin recombinase catalytic domain in the binding of Hin to the hix DNA recombination sites.

The Hin recombinase mediates the site-specific inversion of a segment of the Salmonella chromosome between two flanking 26bp hix DNA recombination sites. Mutations in two amino acid residues, R43 and R69 of the catalytic domain of the Hin recombinase, were identified that can compensate for loss of binding resulting from elimination of certain major and minor groove contacts within the hix recombination sites. With one exception, the R43 and R69 mutants were also able to bind a hix sequence with an additional 4bp added to the centre of the site, unlike wild-type Hin. Purified Hin mutants R43H and R69C had both partial cleavage and inversion activities in vitro while mutants R43L, R43C, R69S, and R69P had no detectable cleavage and inversion activities. These data support a model in which the catalytic domain plays a role in DNA-binding specificity, and suggest that the arginine residues at positions 43 and 69 function to position the Hin recombinase on the DNA for a step in the recombination reaction which occurs either at and/or prior to DNA cleavage.

DNA topology-mediated regulation of transcription initiation from the tandem promoters of the ilvGMEDA operon of Escherichia coli.

It is becoming increasingly clear that the intrinsic and protein-induced topological properties of the DNA helix influence transcriptional efficiency. In this report we describe the properties of two upstream activating regions that influence transcription from the non-overlapping tandem promoters of the ilvGMEDA operon of Escherichia coli. One 20 base-pair region between the promoter sites contains an intrinsic DNA bend that activates transcription from the downstream promoter. The other region contains an integration host factor (IHF) binding site that overlaps the upstream promoter site. IHF binding at this site represses transcription from the upstream promoter and enhances transcription from the downstream promoter. IHF also induces a severe bend in the DNA at its target binding site in the upstream promoter region. The activating property of the 20 base-pair DNA sequence located between the promoters is dependent upon the helical phasing of the sequence-directed DNA bend that it encodes. However, the IHF-mediated activation of transcription is not dependent upon the helical phasing (spatial orientation) of the upstream IHF and downstream promoter sites. The IHF-mediated activation of transcription is also uninfluenced by the presence or absence of the intrinsic DNA bend between its binding site and the downstream promoter site. These results suggest the interesting possibility that IHF activates transcription from the nearby downstream promoter simply by bending the DNA helix in the absence of specific IHF-RNA polymerase or upstream DNA-RNA polymerase interactions.

Carcinosarcoma of the lung with hypertrophic pulmonary osteoarthropathy.

Carcinosarcoma of the lung is a rare malignancy. Endobronchial and parenchymal variants are classically described. Clinicopathological features are often related to anatomical location, as is the case for most lung neoplasms. This case report details the surgical management of a carcinosarcoma in a patient seen with pulmonary osteoarthropathy.

Steady-state and dynamic response properties of the Mandelbaum effect.

The Mandelbaum effect refers to an inadvertent accommodation response to an intervening surface (e.g. a window screen) while attempting to focus a distant object of interest. Previous research has attributed the Mandelbaum effect to a tendency for the accommodation mechanism to preferentially focus the stimulus nearest the dark (resting) focus position. The present study extends this work by examining dynamic accommodation response properties for different distances and stimulus separations. Accommodation measurements of nine young, emmetropic subjects were obtained with an infrared optometer while they viewed superimposed horizontal and vertical square-wave gratings at various dioptric separations. Three subjects demonstrated a strong focussing preference for the target located nearest their dark focus position for all stimulus conditions, while two other subjects showed a similar, but weaker, preference. Conversely, two subjects primarily focussed the nearer of the two targets, and two subjects tended to focus the vertical target (regardless of whether it was near or far, or was closet to the dark focus). For all subjects, accommodation rarely fluctuated from one target to the other, and was seldom focussed midway between the two targets. Our findings indicate that the influence of the dark focus on the Mandelbaum effect varies among individuals, and plays little or no role in some individuals. Thus, accommodation responses to competing stimuli are more complex than previous findings would suggest.

Isolation and characterization of Rhodobacter capsulatus mutants defective in oxygen regulation of the puf operon.

cis-acting mutations that affect regulation of the Rhodobacter capsulatus puf operon by oxygen were isolated by placing the mutagenized puf regulatory region 5' to a promoterless Tn5 neo gene, which encodes resistance to kanamycin (Kmr). R. capsulatus mutants that failed to show wild-type repression of KMr by oxygen were selected and analyzed. Four independent clones contained point mutations, three of which were identical, in a region of dyad symmetry located between puf operon nucleotide positions 177 and 207, approximately 45 base pairs 5' to the site of initiation of puf transcripts. The phenotypic effects of the aerobically selected mutations were duplicated by single and double point mutations introduced site specifically into the region of dyad symmetry by oligonucleotide-directed mutagenesis. Determinations of the bacterial 50% lethal dose of kanamycin, of aminoglycoside phosphotransferase activity in cell sonicates, and of neo-specific mRNA confirmed the diminished responsiveness of the mutants to oxygen and consequently implicated the mutated region in O2-mediated transcriptional regulation.

Macrophage histology in paraffin-embedded multiple sclerosis plaques is demonstrated by the monoclonal pan-macrophage marker HAM-56: correlation with chronicity of the lesion.

Using the monoclonal antibody HAM-56 with the avidin-biotin method on recent or archival paraffin-embedded tissue from multiple sclerosis brains, we have been able to distinguish between acute, chronic active and inactive lesions. HAM-56 stains all macrophages, monocytes and at least some microglia; it is the only pan-macrophage marker to our knowledge that can be satisfactorily used on conventional paraffin sections. A much narrower range of mainly perivascular macrophages in acute plaques of multiple sclerosis is stained with MAC-387, anti-muramidase and anti-alpha1-anti-trypsin. The acute plaques show HAM-56-stained macrophages throughout the lesion, and these macrophages exhibit profiles of phospholipid-rich myelinic bodies, which are also usually stainable with Luxol fast blue. Active ongoing lesions show a rim of macrophages at the edge of the lesion. These macrophages show profiles of large vacuoles, thought to represent the sudanophilic esterified cholesterol formed during demyelination. Inactive cases show none of these features; the few perivascular macrophages present often contain the end product of lipid peroxidation, ceroidlipofuscin.

Perls' ferrocyanide test for iron in the diagnosis of vasculitic neuropathy.

Preliminary observations suggested that arterial and arteriolar necrosis in vasculitis of the peripheral nerve leads to local haemorrhage and subsequent deposition of haemosiderin. This pigment is more readily recognized in the nerve by the sensitive Perls' test for iron than by relying on recognizing its yellow colour. To support the use of iron staining as an index of vasculitis, in addition to necrosis and fibrin deposition, we obtained the following results from nerve biopsy and autopsy nerve specimens: vasculitis confined to PNS = 5/6 iron positive; polyarteritis nodosa and Wegener's granulomatosis = 4/5 iron positive; systemic lupus erythematosus 2/3 iron positive; rheumatoid disease 1/1 iron positive; acute Guillain-Barré syndrome and subacute or chronic demyelinating polyradiculoneuropathy = 12/12 iron negative; and other perivascular inflammation in the PNS (without evidence of vasculitis) = 2/2 iron negative. One case of Churg-Strauss syndrome showed no changes on nerve biopsy. Iron staining was also demonstrated in the kidneys of five or six patients with polyarteritis nodosa.

Pathology, histochemistry and immunocytochemistry of lesions in acute multiple sclerosis.

Twenty cases of acute or early multiple sclerosis have been examined using staining, histochemical or immunocytochemical methods. They had died within 6 months after initial clinical onset (12) or commencement of an "anatomically-remote" acute relapse (8). Plaques in these acute cases showed the following characteristics: lymphocytic perivascular infiltration, plaque hypercellularity, plaque macrophage infiltration and intra-macrophage myelin debris. In most cases of clinical duration of less than 12 weeks, some macrophages showed characteristic formaldehyde-resistant markers for haematogenous macrophages (muramidase, anti-alpha 11-antitrypsin, MAC and HAM56) but, with the exception of the last, these markers subsequently declined indicating a haematogenous origin for macrophages in the early lesion. Lymphocytes were prominent in perivascular (perivenous) regions but, except in one case, were only scanty in or at the demyelinating edge of plaques. Oligodendroglial hyperplasia, indicative of remyelinating activity, was seen at the edge of plaques in one quarter of these acute cases (7 times the rate seen in chronic lesions). Astrocytic activation was not apparent in the earliest stages but was usually seen from about 6 weeks onwards. The conclusion from these observations is that the prime inflammatory process is around blood vessels with usually only scanty initial inflammatory activity in the parenchyma of the brain. Macrophages emigrating from blood vessels digest myelin either as a response to inflammatory damage to the myelin or as a response to activation signals produced in either the perivascular region or plaque.

The cerebral vasculature in dementia pugilistica.

The brains of 22 ex-boxers were examined histologically to determine the frequency of recent or old haemorrhage. Four boxers had died from an acute intracerebral bleed, usually soon after a boxing bout. Seven of the other 18 showed evidence of previous perivascular haemorrhage, as detected by Perls' ferrocyanide test for iron, and a similar number showed minor degrees of meningeal or subpial siderosis, consistent with previous meningeal bleeding; cerebellar siderosis was present in six cases. Seventeen of the 22 boxers showed evidence of recent or past haemorrhage. Control material showed an incidence of 11% for perivascular iron deposition and only 4% for minor degrees of meningeal siderosis.

Structural and functional analysis of transcriptional control of the Rhodobacter capsulatus puf operon.

We report data indicating that the Rhodobacter capsulatus puf operon promoter and the site for its oxygen regulation are located more than 700 base pairs upstream from the previously identified puf genes and have identified the nucleotide sequences that constitute these control signals. A model is proposed in which a polycistronic transcript at least 3.4 kilobases in length is initiated near the O2-regulated promoter and is processed posttranscriptionally by endonucleolytic cleavage at multiple sites, yielding discrete mRNA segments that are degraded at different rates. A newly identified gene (pufQ), which includes a hydrophobic domain having some similarity to domains of the products of the pufL and pufM genes, begins 313 nucleotides into the puf transcript and is located entirely within the most rapidly degraded segment of the transcript. A previously identified puf transcript segment encoding structural proteins for photosynthetic membrane complexes persists after degradation of the most 5' region of the transcript and is itself subject to segmentally specific degradation. Our results suggest a model in which differential expression of the multiple genes encoded by the puf operon is at least in part attributable to major differences in the rates of decay of the various segments of puf mRNA.

The use of colostomy bags for chest tube drainage.

Occasionally patients require long-term chest tube drainage. We describe a method whereby a short chest tube may be drained into a colostomy bag to alleviate the necessity of keeping the patient attached to a bulky drainage system. The colostomy bag allows greater mobility, and the patient can be discharged with the bag.

Coronary arterial dimensions and cell populations in ageing man.

A combined computerised morphometric and immunocytochemical study on the ageing human coronary arterial wall shows that dilatation of the vessel is related to age but is less related to the degree of atherosclerosis. Loss of medial smooth muscle is not related to either atherosclerosis or dilatation of the artery. Occasional cases with severe medial and adventitial infiltrates of inflammatory cells (mainly T-cells and macrophages) do develop marked ectasia, but this arteritis is rather rare and, thus, an infrequent cause of dilatation.

Perivascular iron deposition and other vascular damage in multiple sclerosis.

Evidence of damage to cerebral vein walls was sought in 70 cases of multiple sclerosis. Seventy control cases were also examined. The multiple sclerosis cases showed venous intramural fibrinoid deposition (7%), recent haemorrhages (17%), old haemorrhages revealed by haemosiderin deposition (30%), thrombosis (6%) and thickened veins (19%). In all, 41% of all multiple sclerosis cases showed some evidence of vein damage. Occasional control cases showed haemosiderin deposition in the brain but, unlike the multiple sclerosis cases, these were diffuse and almost entirely related to coexistent cardiovascular or cerebrovascular disease. Haemosiderin deposition was common in the substantia nigra and other pigmented nuclei in all cases. It is concluded that the cerebral vein wall in multiple sclerosis is subject to chronic inflammatory damage, which promotes haemorrhage and increased permeability, and constitutes a form of vasculitis.

Gene organization and structure of the Streptomyces lividans gal operon.

We present the gene organization and DNA sequence of the Streptomyces lividans galactose utilization genes. Complementation of Escherichia coli galE, galT, or galK mutants and DNA sequence analysis were used to demonstrate that the galactose utilization genes are organized within an operon with the gene order galT, galE, and galK. Comparison of the inferred protein sequences for the S. lividans gal gene products to the corresponding E. coli and Saccharomyces carlbergensis sequences identified regions of structural homology within each of the galactose utilization enzymes. Finally, we discuss a potential relationship between the gene organization of the operon and the functional roles of the gal enzymes in cellular metabolism.

Fatigue effects in automated perimetry.

The influence of fatigue and related factors on automated perimetric testing was evaluated in both eyes of sixteen normal observers and sixteen patients with early-to-moderate visual field loss using a Digilab 750 automated perimeter and a customized test procedure. False positive rate, false negative rate, and detection sensitivity at 5,10,15, and 20 degrees eccentricity were measured in 1.5-rain intervals throughout a 21-min visual field examination. Half of the normal observers and patients with visual field loss were given a briefrest(1.5-min) midway through each visual field exam to determine whether this would reduce fatigue effects. Our findings revealed that patients displayed considerably higher average false positive and false negative rates than normal observers. However, neither the patients nor normal observers demonstrated any consistent changes in false positive rate or false negative rate as a function of testing duration. In contrast, both normal observers and patients showed an average decrease in sensitivity as a function of increasing test duration with the magnitude of the time-dependent sensitivity loss becoming greater with increasing stimulus eccentricity. Patients demonstrated a larger time-dependent sensitivity loss than normal observers, averaging ~4dB at 20 degrees eccentricity. The introduction of a brief pause midway through the test procedure appeared to reduce the time-dependent sensitivity loss for the second half of the test procedure, especially for greater eccentricities.