Maternal origin of nucleated erythrocytes in peripheral venous blood of pregnant women

We studied the origin of nucleated red blood cells (NRBC) in peripheral venous blood samples from 40 pregnant women carrying a male fetus, using a technique that allows direct chromosomal analysis by in situ hybridisation on immunologically and morphologically classified cells. Samples from ten nulligravid women were studied as controls. NRBC were enriched by negative magnetic activated cell sorting (miniMACS) using anti-CD45 monoclonal antibody. NRBC were detected by alkaline phosphatase anti-alkaline phosphatase immunostaining using a monoclonal anti-glycophorin A antibody. The origin of the NRBC was determined by fluorescence in situ hybridisation using X and Y specific probes. NRBC were found in 37 of the 40 pregnant women at a range of 1 to 230 per 20 ml of venous blood and in 6 of the 10 controls at a range of 1 to 3 per 20 ml of venous blood. All NRBC detected in the pregnant women were evidently of maternal origin, and in the pregnant women the number of NRBC was significantly higher (P < 0.05) than in the controls. Pregnancy per se seems to induce the appearance of maternal NRBC in the circulation, and it cannot therefore be assumed that NRBC isolated from the maternal blood are of fetal origin on the basis of morphology alone. Discrimination of fetal NRBC must occur for prenatal diagnosis of fetal genetic disorders.     

Meningococcal pilin: a glycoprotein substituted with digalactosyl 2,4-diacetamido-2,4,6-trideoxyhexose

Neisseria meningitidis pili are filamentous protein structures that are essential adhesins in capsulate bacteria. Pili of adhesion variants of meningococcal strain C311 contain glycosyl residues on pilin (PilE), their major structural subunit. Despite the presence of three potential N -linked glycosylation sites, none appears to be occupied in these pilins. Instead, a novel O -linked trisaccharide substituent, not previously found as a constituent of glycoproteins, is present within a peptide spanning amino acid residues 45 to 73 of the PilE molecule. This structure contains a terminal 1-4-linked digalactose moiety covalently linked to a 2,4-diacetamido-2,4,6-trideoxyhexose sugar which is directly attached to pilin. Pilins derived from galactose epimerase ( galE ) mutants lack the digalactosyl moiety, but retain the diacetamidotrideoxyhexose substitution. Both parental (#3) pilins and those derived from a hyper-adherent variant (#16) contained identical sugar substitutions in this region of pilin, and galE mutants of #3 were similar to the parental phenotype in their adherence to host cells. These studies have confirmed our previous observations that meningococcal pili are glycosylated and provided the first structural evidence for the presence of covalently linked carbohydrate on pili. In addition, they have revealed a completely novel protein/saccharide linkage.     

Opc- and pilus-dependent interactions of meningococci with human endothelial cells: molecular mechanisms and modulation by surface polysaccharides

The interplay between four surface-expressed virulence factors of Neisseria meningitidis (pili, Opc, capsule and lipopolysaccharide (LPS)) in host cell adhesion and invasion was examined using derivatives of a serogroup B strain, MC58, created by mutation (capsule, Opc) and selection of variants. To examine the role of Opc and of additional expression of pili, bacteria lacking the expression of Opa proteins were used. The effects of different LPS structures were examined in variants expressing either sialylated (L3 immunotype) or truncated non-sialylated (L8 immuno-type) LPS. Studies showed that (i) pili were essential for meningococcal interactions with host cells in both capsulate and acapsulate bacteria with the sialylated L3 LPS immunotype, (ii) the Opc-mediated invasion of host cells by piliated and non-piliated bacteria was observed only in acapsulate organisms with L8 LPS immunotype, and (iii) expression of pili in Opc-expressing bacteria resulted in increased invasion. Investigations on the mechanisms of cellular invasion indicated that the Opc-mediated invasion was dependent on the presence of serum in the incubation medium and was mediated by serum proteins with arginine-glycine-aspartic acid (RGD) sequence. Cellular invasion in piliated Opc⁺ phenotype also required bridging molecules containing the RGD recognition sequence and appeared to involve the integrin αvβ3 as a target receptor on endothelial cells. These studies extend the previous observations on variants of a serogroup A strain (C751) and show that Opc mediates cellular invasion in distinct meningococcal strains and provide confirmation of its mechanism of action. This is the first investigation that evaluates, using derivatives of a single strain, the interplay between four meningococcal surface virulence factors in host cell invasion.     

Renal arachidonic acid metabolism and cellular changes in the rabbit renal vein constricted kidney: Inflammation as a common process in renal injury models

The process of renal inflammation was examined using the partial renal vein constricted rabbit kidney (RVC) as a model. Forty eight hours of partial renal vein constriction in the rabbit was associated with an increase in prostaglandin (PG) and thromboxane (Tx) production. The perfused RVC kidney showed an enhanced time-dependent increase in PG and Tx production in response to bradykinin stimulation when compared with the unlatered contralateral (CLK) or normal kidney. At 6 hrs of perfusion bradykinin stimulation lateral (CLK) or normal kidney. At 6 hrs of perfusion bradykinin stimulation released 2950±350 ng PGE₂, 61±15 ng TxB₂ from the RVC, and 225±85 ng PGE₂ and undetectable TxB₂ from the CLK. Histological examination of the RVC cortex showed an increase in fibroblast-like cells, a modest increase in the interstitial space and an appearance of macrophages and lymphocytes not seen in the normal of CLK. Endotoxin has been reported to stimulate macrophages in culture to produce PGE₂ and TxB₂. Endotoxin (100 ng)_stimulation of the perfused RVC kidney caused an immediate, followed by a chronically increasing, release of PGs and Tx. Two hours after endotoxin injection 50 ml of effluent fromt the RVC contained 1450±107 ng PGE₂ and 15.0±4.5 ng TxB₂. Other models of renal inflammation (e.g., the hydronephrotic kidney, chronic glomerulonephritis) also show the histological appearance of macrophages. In addition, hydronephrotic kidneys undergo fibroblast proliferation and changes in arachidonic acid metabolism similar to what we observed in the RVC. This work suggests that the inflammatory process (mononuclear cell infiltration), fibroblast-like cell proliferation, and accompanying changes in arachidonate metabolism) is common among different forms of renal injury.     

A survey of dermatophytes isolated from human patients in the United States from 1979 to 1981 with chronological listings of worldwide incidence of five dermatophytes often isolated in the United States

A survey of dermatophytes isolated from patients seeking medical advice was made from 1979 to 1981 in the United States. The survey included 54 locations with data from 40 cities and 2 states. Correlations of these data with that of the other localities of the world were made to illustrate the dynamic epidemiology of several common dermatophytes. The most often isolated dermatophyte in this survey was Trichophyton rubrum having 53.66% of the total for these three calendar years. In a chronological listing of ringworm infections caused by this organism, many areas of the world have reported similar increased incidence of this pathogen. Trichophyton tonsurans was isolated 27.85% of the total. A dramatic increase of this pathogen as a cause of tinea capitis has been observed in most cities of the United States. It has been isolated in 25 different countries of the world. The percentage of isolation of Trichophyton mentagrophytes was 8.56%. This percentage may not be near the true incidence of infection by this dermatophyte because the infections are mild and respond to treatment without the individual seeking medical advice. Since the 1950s the percentage of isolations of the total has dropped for T. mentagrophytes in the United States. Epidermophyton floccosum accounted for 4.36% of the total. In a few areas of the world it causes over 30% of the total of dermatophytoses. Microsporum canis was isolated 3.72% of the total in the United States. It has recently been reported to be the dominant agent of tinea capitis in several South American countries, Tucson, Arizona and Kuwait. Once the dominant pathogen of tinea capitis in children in the United States, it was replaced by Microsporum audouinii before 1960. Today in the United States, M. audouinii only accounts for 0.30% of the total. It is considered eliminated as a pathogen in England. In this survey, isolated less than 1.0% of the total were Microsporum gypseum, Microsporum ferrugineum, Microsporum nanum, Microsporum fulvum and Trichophyton schoenleinii. Trichophyton meginii and Trichophyton terrestre were reported isolated but no numerical data were available.     

Ultrastructure of dynamic and static skeletofusimotor endings in a cat muscle spindle

Summary Endings of four skeletofusimotor axons in a spindle of the cat tenuissimus muscle were examined in semithin (1-m thick) and ultrathin transverse serial sections. Two (dynamic) axons terminated on the nuclear bag₁ intrafusal muscle fiber and on extrafusal fibers of the dark type. Two (static) axons terminated on the nuclear chain intrafusal fibers and extrafusal fibers of the intermediate type. The degree of indentation of axon terminals into the muscle surface, thickness of the sole plate and extent of folding of subjunctional membranes differed among intrafusal and extrafusal terminations of the same axon. Endings of axons on the bag₁ and chain fibers were also morphologically dissimilar. Motor axons may not determine ending morphology. Rather the form and structure of a bag₁ or chain ending may be determined by the type of intrafusal fiber on which the ending lies and the ending's distance from the primary sensory axon.     

Transplantation in miniature swine. XII. N-terminal sequences of class I histocompatibility antigens (SLA) and beta 2-microglobulin

Purified class I histocompatibility antigens (SLA) from three haplotypes were prepared by papain treatment of lymphoid cell membranes obtained from spleens and lymph nodes of miniature swine homozygous at their major histocompatibility complex. Antigens were purified by ion-exchange chromatography followed by gel filtration. Purity was analyzed by SDS-PAGE, and antigenic specificity by inhibition of complement-dependent, alloantiserum-mediated cytotoxicity. The SLA antigens were reduced and alkylated, and the component heavy and light chains were isolated by gel filtration under dissociating conditions. N-terminal amino acid sequences were obtained for SLAaa, SLAcc, and SLAdd heavy chains, as well as for the light chain, beta 2-microglobulin. The swine antigens showed high levels of homology with class I antigens from other animal species. Heterogeneity was observed among the swine haplotypes, and several of the positions at which substitutions were found are apparently invariant in other animal species. In contrast, only minimal sequence heterogeneity was detected within haplotypes, the basis of which may be of relevance to understanding the evolutionary development of these molecules.     

The nuclear matrix: Three-dimensional architecture and protein composition

The structural filament network of the nucleus is prepared while still connected to the cytoskeleton. The relatively gentle procedure removes about 98% of the DNA and at least 86% of the histones. The matrix is bounded by an outer nuclear lamina connected to the cytoskeletal framework, as well as the inner filaments. The filaments range in diameter from 3 to 22 nm, and are organized in a three-dimensional anastomosing network in which nucleoli are enmeshed. The nuclear matrix is separated from the cytoskeletal framework by a double detergent and then partitioned into a chromatin fraction and a matrix fraction by nuclease and high salt. Two-dimensional gel electrophoresis shows that the proteins of the cytoskeleton, chromatin and nuclear matrix are very different. A major protein found in all fractions cofocuses with actin. Vimentin is largely associated with the nuclear matrix, probably as a corona external of filaments.     

Intracellular binding of lead in the kidney: The partial isolation and characterization of postmitochondrial lead binding components

Gel chromatography of kidney postmitochondrial fractions from control rats 2 hr after injection of ²⁰³Pb or after in vitro incubation with ²⁰³Pb disclosed the presence of two fractionated Pb-binding components plus binding in the void volume and total volume regions. The binding of Pb to the two components, with molecular weights of 11,500 and 63,000 daltons, was markedly decreased in Pb-pretreated rats. Sodium dodecyl sulfate-gel electrophoresis and autoradiography showed the presence of one major ²⁰³Pb band with an estimated molecular weight of 60,000 daltons. The 11,500-dalton peak did not incorporate ¹⁴C-leucine nor did concomitant administration of cycloheximide with the ²⁰³Pb inhibit incorporation of ²⁰³Pb activity, suggesting that the component is a preformed constituent of the kidney. In vitro incubation of brain, liver and lung postmitochondrial supernatants with ²⁰³Pb disclosed that these two binding components were also present in brain but not in liver or lung, suggesting a target tissue-specific localization for these Pb-binding macromolecules.